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The Zebrafish Genetics Genomics and Transcriptomics 4th Edition H. William Detrich
The Zebrafish Genetics Genomics and Transcriptomics
4th Edition H. William Detrich Digital Instant Download
Author(s): H. WilliamDetrich, Monte Westerfield and Leonard I. Zon (Eds.)
ISBN(s): 9780128034743, 0128034742
Edition: 4
File Details: PDF, 20.70 MB
Year: 2016
Language: english
Methods in Cell
Biology
The Zebrafish: Genetics,
Genomics, and Transcriptomics
Volume 135
Series Editors
Leslie Wilson
Department of Molecular, Cellular and Developmental Biology
University of California
Santa Barbara, California
Phong Tran
University of Pennsylvania
Philadelphia, USA &
Institut Curie, Paris, France
Methods in Cell
Biology
The Zebrafish: Genetics,
Genomics, and Transcriptomics
Volume 135
Edited by
H. William Detrich, III
Northeastern University Marine Science Center,
Nahant, MA, United States
Monte Westerfield
University of Oregon, Eugene, OR, United States
Leonard I. Zon
Harvard University, Boston, MA, United States
AMSTERDAM • BOSTON • HEIDELBERG • LONDON
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Practitioners and researchers must always rely on their own experience and knowledge in
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and the safety of others, including parties for whom they have a professional responsibility.
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Typeset by TNQ Books and Journals
Len, Monte, and I dedicate the 4th
Edition of Methods in Cell Biology:
The Zebrafish to the postdoctoral fellows and graduate students
who conducted the genetic screens that established the zebrafish as a
preeminent vertebrate model system for analysis of development.
Contributors
J. Ablain
Howard Hughes Medical Institute and Harvard Medical School, Boston, MA,
United States
K. Asakawa
SOKENDAI (The Graduate University for Advanced Studies), Mishima,
Shizuoka, Japan
H. Ata
Mayo Clinic, Rochester, MN, United States
J. Bakkers
Hubrecht Institute and University Medical Centre Utrecht, Utrecht, The
Netherlands
J. Bessa
IBMC-Instituto de Biologia Molecular e Celular, Porto, Portugal; Universidade do
Porto, Porto, Portugal
Y.M. Bradford
University of Oregon, Eugene, OR, United States
B.R. Cairns
University of Utah School of Medicine, Salt Lake City, UT, United States
W. Chen
Vanderbilt University School of Medicine, Nashville, TN, United States
J. Cibelli
Michigan State University, East Lansing, MI, United States; BIONAND,
Andalucı́a, Spain
K.J. Clark
Mayo Clinic, Rochester, MN, United States
P. Coucke
Ghent University, Ghent, Belgium
V.T. Cunliffe
University of Sheffield, Sheffield, United Kingdom
F. Del Bene
PSL Research University, Paris, France
xvii
A. De Paepe
Ghent University, Ghent, Belgium
F. De Santis
PSL Research University, Paris, France
V. Di Donato
PSL Research University, Paris, France
A. Eagle
University of Oregon, Eugene, OR, United States
S.C. Ekker
Mayo Clinic, Rochester, MN, United States
T. Erickson
Oregon Health & Science University, Portland, OR, United States
T. Evans
Weill Cornell Medical College, New York, NY, United States
D. Fashena
University of Oregon, Eugene, OR, United States
A. Felker
University of Zürich, Zürich, Switzerland
A. Fernández-Miñán
Centro Andaluz de Biologı́a del Desarrollo (CABD), Consejo Superior de
Investigaciones Cientı́ficas/Universidad Pablo de Olavide/Junta de Andalucı́a,
Sevilla, Spain
K. Frazer
University of Oregon, Eugene, OR, United States
A. Ghosha
Carnegie Institution for Science, Baltimore, MD, United States
M.G. Goll
Memorial Sloan Kettering Cancer Center, New York, NY, United States
J.L. Gómez-Skarmeta
Centro Andaluz de Biologı́a del Desarrollo (CABD), Consejo Superior de
Investigaciones Cientı́ficas/Universidad Pablo de Olavide/Junta de Andalucı́a,
Sevilla, Spain
a
Current address: Tata Institute of Fundamental Research, Mumbai, India
xviii Contributors
D.J. Grunwald
University of Utah, Salt Lake City, UT, United States
M.E. Halpern
Carnegie Institution for Science, Baltimore, MD, United States
J.K. Heath
Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia
K. Hoshijima
University of Utah, Salt Lake City, UT, United States
D.G. Howe
University of Oregon, Eugene, OR, United States
H. Huang
University of California Los Angeles, Los Angeles, CA, United States
J.P. Junker
Hubrecht Institute and University Medical Centre Utrecht, Utrecht, The
Netherlands
M.J. Jurynec
University of Utah, Salt Lake City, UT, United States
P. Kalita
University of Oregon, Eugene, OR, United States
K. Kawakami
SOKENDAI (The Graduate University for Advanced Studies), Mishima, Shizuoka,
Japan
M.C. Keightley
Australian Regenerative Medicine Institute, Clayton, VIC, Australia; Monash
University, Clayton, VIC, Australia
F. Kruse
Hubrecht Institute and University Medical Centre Utrecht, Utrecht,
The Netherlands
C. Lawrence
Boston Children’s Hospital, Boston, MA, United States
S. Lefever
Ghent University, Ghent, Belgium
C. Li
Memorial Sloan Kettering Cancer Center, New York, NY, United States; Weill
Cornell Graduate School of Medical Sciences, New York, NY, United States
Contributors xix
G.J. Lieschke
Australian Regenerative Medicine Institute, Clayton, VIC, Australia; Monash
University, Clayton, VIC, Australia
S. Lin
University of California Los Angeles, Los Angeles, CA, United States
C.G. Love
Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia;
University of Melbourne, Parkville, VIC, Australia
L.A. Maddison
Vanderbilt University School of Medicine, Nashville, TN, United States
P. Mani
University of Oregon, Eugene, OR, United States
S. Markmiller
University of California San Diego, La Jolla, CA, United States
R. Martin
University of Oregon, Eugene, OR, United States
S. Masuda
Tokyo Institute of Technology, Yokohama, Japan
A.C. Miller
University of Oregon, Eugene, OR, United States
C.B. Moens
Fred Hutchinson Cancer Research Center, Seattle, WA, United States
C. Mosimann
University of Zürich, Zürich, Switzerland
S.T. Moxon
University of Oregon, Eugene, OR, United States
M.C. Mullins
University of Pennsylvania Perelman School of Medicine, Philadelphia, PA,
United States
P.J. Murphy
University of Utah School of Medicine, Salt Lake City, UT,
United States
K.N. Murray
University of Oregon, Eugene, OR, United States
xx Contributors
A. Muto
SOKENDAI (The Graduate University for Advanced Studies), Mishima,
Shizuoka, Japan
T. Nicolson
Oregon Health & Science University, Portland, OR, United States
C.J. Ott
Dana Farber Cancer Institute, Harvard Stem Cell Institute, Boston, MA,
United States
H. Paddock
University of Oregon, Eugene, OR, United States
F. Pelegri
University of WisconsineMadison, Madison, WI, United States
C. Pich
University of Oregon, Eugene, OR, United States
S. Prukudom
Kasetsart University, Bangkok, Thailand; Center of Excellence on Agricultural
Biotechnology: (AG-BIO/PERDO-CHE), Bangkok, Thailand
S. Ramachandran
University of Oregon, Eugene, OR, United States
J.E.J. Rasko
Centenary Institute, Camperdown, NSW, Australia; University of Sydney,
Sydney, NSW, Australia; Royal Prince Alfred Hospital, Newtown, NSW,
Australia
M.P. Rossmann
Harvard University, Harvard, Cambridge, MA, United States
L. Ruzicka
University of Oregon, Eugene, OR, United States
K. Schaper
University of Oregon, Eugene, OR, United States
A.N. Shah
Fred Hutchinson Cancer Research Center, Seattle, WA, United States
X. Shao
University of Oregon, Eugene, OR, United States
A. Singer
University of Oregon, Eugene, OR, United States
Contributors xxi
K. Siripattarapravat
Kasetsart University, Bangkok, Thailand; Center of Excellence on Agricultural
Biotechnology: (AG-BIO/PERDO-CHE), Bangkok, Thailand
F. Speleman
Ghent University, Ghent, Belgium
M. Superdock
Boston Children’s Hospital, Boston, MA, United States; Dana Farber Cancer
Institute, Harvard Stem Cell Institute, Boston, MA, United States; Harvard
Medical School and Howard Hughes Medical Institute, Boston, MA,
United States
M. Tanaka
Tokyo Institute of Technology, Yokohama, Japan
J.J. Tena
Centro Andaluz de Biologı́a del Desarrollo (CABD), Consejo Superior de
Investigaciones Cientı́ficas/Universidad Pablo de Olavide/Junta de Andalucı́a,
Sevilla, Spain
S. Toro
University of Oregon, Eugene, OR, United States
A. van Oudenaarden
Hubrecht Institute and University Medical Centre Utrecht, Utrecht,
The Netherlands
J. Vandesompele
Ghent University, Ghent, Belgium
S. Vanhauwaert
Ghent University, Ghent, Belgium
C. Van Slyke
University of Oregon, Eugene, OR, United States
Z.M. Varga
University of Oregon, Eugene, OR, United States
H. Wada
SOKENDAI (The Graduate University for Advanced Studies), Mishima,
Shizuoka, Japan
M. Westerfield
University of Oregon, Eugene, OR, United States
xxii Contributors
A. Willaert
Ghent University, Ghent, Belgium
S. Yang
Boston Children’s Hospital, Boston, MA, United States; Dana Farber Cancer
Institute, Harvard Stem Cell Institute, Boston, MA, United States; Harvard
Medical School and Howard Hughes Medical Institute, Boston, MA, United
States
L. Yin
Vanderbilt University School of Medicine, Nashville, TN, United States
B. Zhang
Peking University, Beijing, People’s Republic of China
Y. Zhang
Peking University Shenzhen Graduate School, Shenzhen, China; University of
California Los Angeles, Los Angeles, CA, United States
Y. Zhou
Boston Children’s Hospital, Boston, MA, United States; Dana Farber Cancer
Institute, Harvard Stem Cell Institute, Boston, MA, United States; Harvard
Medical School and Howard Hughes Medical Institute, Boston, MA, United
States; Harvard University, Harvard, Cambridge, MA, United States
L.I. Zon
Boston Children’s Hospital, Boston, MA, United States; Dana Farber Cancer
Institute, Harvard Stem Cell Institute, Boston, MA, United States; Harvard
Medical School and Howard Hughes Medical Institute, Boston, MA, United
States; Harvard University, Harvard, Cambridge, MA, United States
Contributors xxiii
Preface
Len, Monte, and I are pleased to introduce the fourth edition of Methods in Cell
Biology: The Zebrafish. The advantages of the zebrafish, Danio rerio, are numerous,
including its short generation time and high fecundity, external fertilization, and the
optical transparency of the embryo. The ease of conducting forward genetic screens
in the zebrafish, based on the pioneering work of George Streisinger, culminated in
screens from the laboratories of Wolfgang Driever, Mark C. Fishman, and Christiane
Nüsslein-Volhard, published in a seminal volume of Development (volume 123,
December 1, 1996) that described a “candy store” of mutants whose phenotypes
spanned the gamut of developmental processes and mechanisms. Life for geneticists
who study vertebrate development became really fine.
Statistics derived from ZFIN (the Zebrafish Model Organism Database; http://zfin.
org) illustrate the dramatic growth of research involving zebrafish. The zebrafish
genome has been sequenced, and as of 2014, more than 25,000 genes have been
placed on the assembly. Greater than 15,500 of these genes have been established
as orthologs of human genes. The zebrafish community has grown from w1,400 re-
searchers in 190 laboratories as of 1998 to w7,000 in 930 laboratories in 2014. The
annual number of publications based on the zebrafish has risen from 1,913 to 21,995
in the same timeframe. Clearly, the zebrafish has arrived as a vertebrate biomedical
model system par excellence.
When we published the first edition (volumes 59 and 60) in 1998, our goal was to
encourage biologists to adopt the zebrafish as a genetically tractable model organism
for studying biological phenomena from the cellular through the organismal. Our
goal today remains unchanged, but the range of subjects and the suite of methods
have expanded rapidly and significantly in sophistication over the years. With the
second and third editions of MCB: The Zebrafish (volumes 76 and 77 in 2004;
volumes 100, 101, 104, and 105 in 2010e11), we documented this extraordinary
growth, again relying on the excellent chapters contributed by our generous col-
leagues in the zebrafish research community.
When Len, Monte, and I began planning the fourth edition, we found that the
zebrafish community had once more developed and refined novel experimental
systems and technologies to tackle challenging biological problems across the
spectrum of the biosciences. We present these methods following the organizational
structure of the third edition, with volumes devoted to Cellular and Developmental
Biology, to Genetics, Genomics, and Transcriptomics, and to Disease Models and
Chemical Screens. Here we introduce the third volume, Genetics, Genomics, and
Transcriptomics.
Genetics, Genomics, and Transcriptomics is divided into five sections that cover
genetic and genomics techniques. Part 1 covers forward and reverse genetics in nine
chapters, many of which employ the revolutionary CRISPR/Cas9 technology in
novel ways. Precision editing of the zebrafish genome through homologous recom-
bination has now become a reality. Part 2 contains five chapters that describe
xxv
advances in transgenesis and functional genomics approaches. Spatially resolved
transcriptomics at the organismal level, cell type-specific transcriptomics, and the
important companion technology, RT-qPCR, are presented in Part 3. We devote
Part 4 to five chapters on the emerging analysis of epigenetic regulation of gene
expression in the zebrafish. Part 5 concludes the volume with four important chap-
ters on zebrafish husbandry, health monitoring, disease prevention, and information
technology.
We anticipate that you, our readership, will apply these methods successfully in
your own zebrafish research programs and will develop your own technical advances
that may be considered for a future edition of Methods in Cell Biology: The Zebra-
fish. The zebrafish is a remarkable experimental systemdthe preeminent vertebrate
model for mechanistic studies of cellular and developmental processes in vivo.
We thank the series editors, Leslie Wilson and Phong Tran, and the staff of
Elsevier/Academic Press, especially Zoe Kruze and Sarah Lay, for their enthusiastic
support of our fourth edition. Their help, patience, and encouragement are pro-
foundly appreciated.
H. William Detrich, III
Monte Westerfield
Leonard I. Zon
xxvi Preface
Multiplex conditional
mutagenesis in zebrafish
using the CRISPR/Cas
system
1
L. Yina
, L.A. Maddisona
, W. Chen1
Vanderbilt University School of Medicine, Nashville, TN, United States
1
Corresponding author: E-mail: wenbiao.chen@vanderbilt.edu
CHAPTER OUTLINE
Introduction................................................................................................................ 4
1. Methods ................................................................................................................ 5
1.1 Assembly of U6-Based sgRNA Transgenic Constructs................................. 5
1.2 Construction of Cas9 Expression Vectors ................................................... 9
1.3 Screening and Evaluation of Stable sgRNA or Cas9 Transgenic Fish ............ 9
2. Discussion........................................................................................................... 14
Summary .................................................................................................................. 14
Acknowledgments..................................................................................................... 15
References ............................................................................................................... 15
Abstract
The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-asso-
ciated protein (Cas) system is a powerful tool for genome editing in numerous organisms.
However, the system is typically used for gene editing throughout the entire organism.
Tissue and temporal specific mutagenesis is often desirable to determine gene function in
a specific stage or tissue and to bypass undesired consequences of global mutations. We
have developed the CRISPR/Cas system for conditional mutagenesis in transgenic
zebrafish using tissue-specific and/or inducible expression of Cas9 and U6-driven
expression of sgRNA. To allow mutagenesis of multiple targets, we have isolated four
distinct U6 promoters and designed Golden Gate vectors to easily assemble transgenes
with multiple sgRNAs. We provide experimental details on the reagents and applications
for multiplex conditional mutagenesis in zebrafish.
CHAPTER
a
These authors contributed equally.
Methods in Cell Biology, Volume 135, ISSN 0091-679X, http://dx.doi.org/10.1016/bs.mcb.2016.04.018
© 2016 Elsevier Inc. All rights reserved.
3
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169
charge and brought with him forty-seven men under James Andrus
with wagons and supplies for an extended trip designed to drive the
Navajos across the Colorado River. When they arrived at Pipe Springs,
the snow was two feet deep and no trace of the sheep or men could
be found. On January 18, they came upon the tracks of two Paiute
Indians following a large steer, tracked them until sundown, and
captured the Indians in the act of killing the beef.
After questioning and torture, hanging by the heels and twisting of
thumbs, one of the Indians admitted that he had dreamed that
Navajos had been there and then revealed the whereabouts of a
camp of Indians about ten miles out. A detachment was sent and
found that it had been moved another five miles. The militia overtook
the camp about sunrise on January 20, killing two Indians and
capturing five.
Third degree methods elicited information about the killing of
Whitmore and McIntyre. The captives led another detachment to the
scene of the killings, where the posse crisscrossed the area on
horseback, uncovering the arm of one of the victims in the deep
snow. Both bodies had bullet wounds and were riddled with arrows.
They had been killed on January 10.
A wagon was sent after the bodies. While the men were recovering
the remains the other detachment with the five Indian prisoners
arrived. These had in their possession much of the clothing and
personal effects of the murdered men. The evidence of guilt seemed
conclusive, so the Indians were turned loose and shot as they
attempted to run. The Navajos who probably assisted in the killing
escaped. The sheep could not be found and it was assumed
the Navajos had taken them across the Colorado River. As
pursuit was impossible because of the deep snow the party returned
home. Charles L. Walker of St. George records in his diary:
They were brought home in a wagon load of snow, frozen stiff and
in a good state of preservation. I, with others, washed them and
pulled out the arrow points from their bodies and dressed them in
their burial robes. Also went to the funeral, which was attended by
a large concourse of people.
[70]
On February 19, 1866, two days after Erastus Snow was elected
Brigadier General, Peter Shurtz, who had built a station at Paria and
had kept about twenty Indians around him all winter, reported that
he had lost his cattle and wished to move into the settlements. He
also reported Navajos camped on Paria River about eight miles below
his ranch where the Ute trail reached the stream.
Further information indicated that the Navajos were concentrated
east of the Colorado at Cottonwood, intending to raid Kane County in
force and that Captain James Andrus with thirty men had gone to
Paria to get Peter Shurtz and his family and to reconnoiter. No report
of this expedition is available, but a letter written by L. W. Roundy
from Kanab on March 9, 1866, tells that Andrus had left Paria
fourteen days earlier headed for an Indian camp twelve miles south.
[71]
At Kanab, three Indians had attempted to kill Oren Clark in the
bottoms near the fort and had started to drive off the livestock. Four
men from the fort rushed in pursuit and after dark recovered about
thirty head of cattle, but the Indians escaped with about an equal
number.
The Indian menace was so serious by this time that Erastus Snow
ordered all stock in the region south of St. George and the Virgin
River as far east as Kanab removed to the north and west of the lines
of settlements so that it would be easier to ward off Navajo attacks.
This was a difficult task because the grazing was poor around the
settlements and the mountains to the northwest were already filled
with livestock.
The threat from the Utes in upper Sevier Valley also became acute.
Menacing behavior of the Indians in this area and in the Kanab region
led to an order from Utah headquarters to General Erastus Snow
(March 15) to send a company of men from Beaver and Iron counties
over to the Sevier River to build and man an outpost between
170
Circleville and Panguitch. A company of 76 men led by Captain Silas
S. Smith served here from March 21 to November 30, 1866. They
established Fort Sanford about ten miles north of Panguitch and
assisted settlers at Circleville to move to safety. At Panguitch,
they helped the settlers transform the town into a fort.
In the meantime, gathering the livestock from the exposed range was
proceeding slowly. A party sent out from Rockville in April to round up
the stock in the vicinity of Maxwell’s Ranch, found the bodies of two
men, a woman, and an Indian, killed a few days before. When the
bodies were brought in to Grafton, it was ascertained that they were
young Robert Berry, his wife, Isabel, and his brother, Joe, who were
coming home to Berryville in Long Valley via the Dixie settlements
and the Arizona Strip (a roundabout way, but the only wagon route at
the time). They had left the Maxwell Ranch on Short Creek on April 2,
1866 when some Indians (presumably Paiutes), ambushed them.
According to verbal reports, as related by Mrs. John Dennett of
Rockville (then a girl living in Long Valley and who pieced her story
from Indian and white sources), the Berrys fought for their lives. The
Indians shot one of the horses, rendering the wagon useless. In the
fighting, one Indian was shot. Joe Berry loosened the other horse
and tried to escape but was killed in so doing. The Indians closed in
and captured Robert and his wife. They tied Robert to a wheel where
he was forced to watch them torture Isabel, who was an expectant
mother. They shot arrows into her and laughed at her as she tried to
pull them out. Then they shot him full of arrows. Mrs. Dennett said
her father always felt that the Berrys had been killed in revenge for
some Indians slain by Long Valley men who had found them roasting
a beef. At that time three were slain: an Indian, a squaw and a
papoose.
When the Berry tragedy was reported in St. George, orders were
issued forbidding travel unless in groups large enough to provide
adequate safeguards. This led to the declaration of martial law, May
2, 1866, and to the issuance of instructions to concentrate the
171
settlers in fortified places of at least 150 men. Patrols were ordered
out in various directions, especially across the trails used by the
Navajos in raiding the Mormon country and in contacting the
rampaging Utes of Sevier County.
When Silas S. Smith, stationed on the Sevier, heard of the Berry
massacre, he found that the Paiute chief at Panguitch had known
about it for five days without reporting it to him. Smith at once
ordered pickets to bring in all passing Indians for questioning.
Friendly Indians responded willingly enough, but when two strange
Indians refused, a skirmish resulted in which one was killed and the
other wounded.
Smith decided to disarm the local Indians and surrounded one of
their camps near Panguitch one morning before daylight and took
their arms. Two visiting Indians were missing from the camp so he
kept a guard awaiting their arrival. When they came, they showed
fight. One of them was killed, whereupon the other surrendered. The
next day Smith surrounded another camp soon after sunrise,
but the natives had already fled. However, in the ensuing
melee two more Indians were killed. The arms taken from them
included several guns, many new arrows, and a peck of new arrow
heads. Some escaped to Panguitch Lake and spread the alarm among
the Indians there.
General Snow had a number of chiefs from Panguitch, Parowan, and
Red Creek brought to Parowan for conference. He tried to pacify
them with arguments and presents but insisted that they must not
have arms or ammunition and must have passes in order to travel
through the Mormon settlements. This aroused some resentment, but
on April 25, 1866, they agreed to leave their weapons at Parowan as
a token of friendship. Some of the Indians reported gunfire around
Upper Kanab where Col. W. B. Maxwell was on lookout for Navajos.
With the declaration of martial law and the order to concentrate
settlers in large towns, the outlying ranchers and people from the
smaller villages began to move into Toquerville, Virgin and Rockville.
172
In June, General Snow decided to abandon Long Valley. Mrs. John
Dennett, who made the trek as a girl, recalled the line of wagons
leaving Long Valley with armed guards in front and rear. While
crossing the sand hills between there and Kanab, a small boy was run
over and killed. A halt was made while the child was buried in the
sand, but the exigencies of the situation forbade longer delay and the
weeping mother was hastily torn from the fresh grave.
The settlers’ train passed Kanab to the left and pushed on toward
Pipe Springs. Near the mouth of Cottonwood Canyon (June 27) they
ran into an ambush of Indians who, for some unexplained reason,
failed to attack. J. M. Higbee reports
[72]
that they called to the
Indians to come in and talk or be shot. They came in and talked.
According to Mrs. Dennett, there were seven or nine Indians taken
into Pipe Springs for a council of war. The wagons were driven into a
large circle, as was customary in times of danger, the Indians inside
the circle in the center of the group of whites. Higbee says the
Indians were told that if any more were found along the route of the
caravan they would be shot. Mrs. Dennett adds that some of the
Indians had guns and clothes belonging to the Berry boys, which
greatly enraged a brother of the dead men, who pleaded to be
allowed to revenge his kin. After this, no more Indians were seen on
the trip.
In the late summer of 1866, Captain James Andrus
[73]
was
ordered to investigate Indian routes crossing the Colorado
River in the rough country between the Kaibab and the mouth of the
Green River. A group of men was mustered into service from the
Virgin River settlements at Gould’s Ranch, twenty-six miles east of St.
George, August 16, and moved on to Pipe Springs two days later,
where final preparations were made. On the 21st, forty-six mounted
men, each equipped with a rifle and two pistols, and with a pack
horse bearing forty days’ rations for each pair, started northeast
toward the rough country. They went via abandoned Kanab and
Scutumpah to the Paria River six miles above Paria settlement where
they met another contingent of their party. Two days later, Joseph
Fish with eighteen men arrived from Iron county. They located the
Ute trail which passed down the Paria to the Colorado. Elijah Averett,
sent back with some of the surplus animals, was killed by Indians in
the hills west of Paria.
On August 29, the main party went northeast through the hills south
of the Aquarius Plateau into a valley where they found wild potatoes
growing (hence named Potato Valley, now Escalante Valley). They
climbed the Plateau and looked off into the wild country stretching to
the mouth of the Green River. Convinced that there was no use in
going farther, they retraced their march on September 2, traveled to
the northwest corner of the plateau, descended to the Sevier River
Valley and reached Circleville. They had been pathbreakers from Paria
to this point. From here they returned via Parowan and Cedar City.
The settlements were now prepared for attack. An Indian raid was
made on John D. Lee’s ranch near Beaver on October 23, 1866, and
in November, General Snow learned that the Navajos were
concentrating east of the Colorado for new raids on Kane County.
Soon a friendly Paiute reported that the Navajos were nearing Pipe
Springs.
The crops planted in Long Valley had been left in the care of friendly
Paiutes when the settlers left. In the fall, the Berry boys and others
went back to harvest the best crop that had yet been grown there. It
took several trips to haul the produce to the Dixie settlements. During
their last trip, Snow received a report of an attack of sixteen Navajos
on three white men at Maxwell’s Ranch, in which Enoch Dodge was
wounded. Snow sent men to Long Valley and instructed A. P. Winsor
to throw an intercepting force between the settlements and the fords
of the Colorado, to recover lost stock and find out whether the
raiders were Navajos or Paiutes. He was promised that other men
would be held in readiness if needed.
While this force was on the road, the Long Valley party started home
with a wagon train. On October 31, when the teams were spread out
173
doubling up Elephant hall, about nine miles south of Mt. Carmel,
Indians attacked near the summit and shot Hyrum Stevens.
The pioneers abandoned the train and left everything in the
hands of the Indians. Stevens was taken with the others on
horseback (with a man behind to hold him in place) around the head
of Zion Canyon on a three-day trip over the Old Indian Trail and down
over Kolob to Virgin. He survived the ordeal and returned to his home
at Rockville, where he lived to a ripe old age.
When a rescue party under Captain Sixtus E. Johnson arrived a week
later, November 5, they found the wagons unattended, tongues
broken and contents scattered. The Indians had taken five yoke of
oxen, eleven horses and everything they could carry, including
harness, flour and wheat. Four Paiutes, however, had pursued the
Navajos and recaptured the cattle and harness.
Finding the teamsters gone, Johnson gathered up the livestock that
had scattered back along the way to Long Valley. Then a second
rescue party under Major Russell from Rockville arrived with the
Paiutes who had retrieved the harness and cattle. They took the
caravan into Virgin, arriving November 11.
On November 26, Major John Steele reported signal fires on the
mountain south of Virgin City and General Snow issued an order to
establish posts at the mouth of (Black) Rock Canyon sixteen miles
southeast of St. George and near Gould’s Ranch, eight miles south of
Virgin City.
[74]
The men at these posts were to serve as guards as
well as herders of livestock and were to build stone quarters; the
“house to be covered with stone flagging or earth in a manner that it
cannot be fired from the outside, with but one door and that heavy
and strongly barred, so that one or two men, well armed, may
defend themselves against any number of Indians.”
Despite these precautions, the Navajos scattered in small bands,
easily passed through the military posts, and hid in the mountains
north of St. George. On the evening of December 28, word reached
174
Harrisburg from local Paiutes that some Navajos had killed and dried
three beeves between Grapevine Springs and Toquerville. Captain J.
D. L. Pearce, with fifteen men from Washington, at once took up their
trail along Harrisburg Creek toward Pine Valley Mountain but failed to
overtake them.
In the meantime, on December 28, near Pine Valley, Cyrus Hancock
saw three Indians skulking on the range and called to them. The
Indians proved hostile and tried to capture him. One seized his
horse’s bit and another tried to shoot him with an arrow. He slid off
his horse and ran toward Pine Valley, the Indians in pursuit. One of
them shot him in the arm with an arrow. He stumbled and fell as they
yelled in triumph, but he regained his footing and outran them into
the valley. These Indians were thought to have been hiding around
the town for two or three days, quietly gathering stock. As
soon as discovered, they left with about thirty horses and
passed down the Black Ridge between St. George and Middleton on
the night of December 28, gathering more horses at both places and
hastening southwest via Fort Pearce Wash.
Col. D. D. McArthur immediately ordered out all available cavalry in
pursuit of the thieves, who had an entire day’s start. An expedition of
thirty men headed by Lt. Copelan followed the Indian trail from the
Washington Fields past Fort Pearce, through Black Rock Canyon and
out toward Pipe Springs where it met another detachment returning
from an Indian encounter.
Captains J. D. L. Pearce and James Andrus were at Harrisburg on the
evening of December 29, when an express carrying instructions to
Colonel Winsor at Rockville arrived. Upon reading the instructions
they started for Rockville and arrived at dawn. Thirty men gathered
and pushed on to Maxwell’s ranch where they arrived that evening.
After resting an hour, they hastened on to Cedar Ridge and five miles
southeast of Pipe Springs. Sixtus E. Johnson spotted the smoke of
Indian fires curling up in the distance, about half a mile from the
place where Whitmore and McIntyre had been killed. The men
175
slipped into a wash and kept out of sight until within gunshot of the
Indians, when they made a dash to get between them and their
horses. Firing opened and the Indians took to the rocks. The skirmish
lasted nearly an hour and covered a rough area half a mile wide and
three miles long. The thirteen Navajos in the band refused to yield
even when cornered, and several died fighting. During the fray an
arrow aimed at Captain Andrus struck his horse in the forehead,
saving the rider. One mortally wounded Indian continued to shoot
until he fainted. Another, wounded in both legs, fired until his arrows
were spent and then kept twanging his bow as if shooting as long as
the fray lasted.
When all was quiet the whites gathered together and found that none
was injured. Two Indians who had escaped came out on a hill some
distance away where they felt safe and slapped their seats in derision
and yelled “Squaw! Squaw!” in defiance. A man named Warren, from
Pine Valley, who had an extra long range breechloading gun took a
chance shot and brought one of them down. The other fled.
Of the thirteen Indians, four were killed, seven wounded and two
escaped, only one on horseback. Three horses were lost, but the
balance and the thirteen cattle were recovered and brought back.
Copelan’s party returned on January 1, 1867 but Pearce and Andrus
tarried two or three days longer. General Snow was in Salt Lake City
at the time and his responsibility fell largely upon Captain J. D. L.
Pearce and Adjutant Henry Eyring, his assistant.
The concentration in the larger towns and the military control of the
movements of people in the region tended to reduce the danger to
the settlers. Tension with the local Paiute Indians was
gradually eased, although the Navajo raids continued for
several years. Jacob Hamblin, Utah’s “Leatherstocking,” played an
important role in quieting the Paiutes. In the fall of 1867, he was
instructed to keep in touch with the Indians and do his best to pacify
them. He went to Kanab, where he helped them plant corn and
vegetables and had peace parleys with them, urging the Paiutes to
cooperate in preventing Navajo raids by watching the fords of the
Colorado and the trails leading to the settlements.
In November, 1868, a band of about thirty Navajos crossed the river
on foot on a marauding expedition. They divided into squads of two
or three and worked at night in different quarters so rapidly as to
baffle the pickets. They got away with some stock, although twenty-
seven horses were recovered from them on the 25th at Black
Canyon, by Andrus and his command.
Notice of their presence came on November 22 from Henry Jennings
to Erastus Snow at St. George. The next day local Indians reported
tracks around St. George, and General Snow ordered the livestock
along the Virgin gathered together and herded under armed guard.
He placed pickets along the river for fifteen miles and sent Col. J. D.
L. Pearce with a company of cavalry to guard the rough country
passes from Black Rock Canyon (25 miles southeast of St. George) to
Pipe Springs. Two days later, word came from Washington that the
Navajos had made off with a band of horses via Black Rock Canyon.
On the night of the 26th, a party of these Indians with about twenty
horses eluded the guards not far from Pipe Springs and made their
way eastward. A detachment under Captain Willis Copelan started in
pursuit. He chased the Navajos and was about to overtake them, but
before he attacked, some friendly Paiutes encountered the Navajos,
gave battle and killed two. They recovered the horses and willingly
turned them over to Copelan on his arrival. The Paiutes were
rewarded with suitable presents.
By December 1, 1868 Pearce concluded that the Navajos had
decamped, and started home, moving from Pipe Springs to Cedar
Ridge. On that same day, however, Erastus Snow received word from
J. W. Young on the Muddy River in Nevada, sixty miles below St.
George, that the Navajos had run off with eighteen horses and
mules. Snow sent word to Pearce to be on his guard. A posse of
whites and Paiutes set out in pursuit from Mesquite. The Indians
overtook the Navajos and recovered eleven of the horses.
176
The messenger carrying this news reached Col. Pearce at Cedar
Ridge at 4 a.m., December 2, and at daybreak scouts were sent out.
Captain Freeman found their trail and started after them with several
men, being joined by Captain Copelan. They sighted the Navajos’
dust, but could not overtake them and the chase had to be
abandoned.
A raid in February 1869 caused such concern that another
expedition (February 25 to March 12) of thirty-six men, under
the leadership of Captain Willis Copelan was sent out to deal with it.
[75]
As usual, the Navajos struck swiftly and fled before the expedition
arrived. At Pipe Springs, Copelan watched the passes, hunted the
surrounding region for the raiders and found they had gone east. On
March 1, with twenty men he started in pursuit of the raiders. About
eight miles out he struck a trail where the Indians had been driving
about fifty head of cattle. During the next five days he followed the
trail around the north end of Buckskin Mountain (Kaibab) across Paria
and Warm Springs Creek to the old Ute ford on the Colorado River.
Finding the quarry had escaped, he returned home, arriving at St.
George March 12.
During the fall, fresh raids by the Navajos created yet another scare.
A band raided settlements north of St. George and drove off stock.
This time Colonel James Andrus was detailed to lead a foray against
the marauders. He started up the Virgin River gathering fourteen
recruits. Then he went to Pipe Springs where he received word that
another band of Navajos had raided near Pinto. He hastened toward
Pinto to intercept them, passing via Kanab and Scutumpah. Near
Paria, he found a trail where some Navajos had escaped with an
estimated eighty head of livestock. Here Andrus learned that the
Paiutes had attacked and wounded a Navajo in a running fight, and
that other raiders were on the way back from Pinto.
Andrus and his men waited until November 10 and finding no signs of
the Indians, started home. The detachment had not gone far when
they encountered a fresh Navajo trail made by an estimated twelve
177
horses and two men. They caught up with the Indians early the next
day just as they were passing into a narrow gorge of the Paria
canyon. There were actually eight Indians with twelve horses,
traveling leisurely. Under the detachment’s fire, two Navajos fell; the
rest disappeared into the narrows. A few minutes later they re-
appeared on the canyon cliffs on both sides of Andrus’ force. Bullets
from the Indian rifles soon convinced Andrus that discretion was the
better part of valor and he retired.
The Navajos were adroit raiders. In rounding up stock they would
often camouflage themselves with bush foliage, crawling past the
unsuspecting guards to stampede the herd. Or they would skin a
young steer, leaving hoofs and horns in place and throw the hide over
a brace of Indians, who would steal to the corral under cover of
darkness, let down the bars, and quietly drive the stock away.
These raids were costly. Not only did the settlers live in constant fear,
but a heavy toll of livestock, estimated in 1869 at 1200 horses
and cattle, was taken. Men had to be continually on the alert
and peaceful pursuits were interrupted to furnish posses to chase the
Indians. When Major J. W. Powell of the U. S. Geological Survey was
exploring the Kanab region in 1870, he expressed grave concern
about the losses the Mormon settlers were suffering because of the
raids.
[76]
In October, Jacob Hamblin decided to accompany Major
Powell on a peace mission to the Navajos when the latter was leaving
to return to Washington, DC.
[77]
They reached Fort Defiance in
eastern Arizona at a time when 6,000 Navajos were gathered there
for their annual allotments from the Federal government.
All the Navajo chiefs but one were present and met in council to
consider Hamblin’s proposal. Powell introduced Hamblin by saying
that he represented the Mormons from the other side of the Colorado
River who were helping to pay the taxes from which the annual
allotments to the Navajos were made. Hamblin, in turn, pointed to
the disastrous consequences of the war and the advantages of peace.
178
Through war the Navajos had lost twenty or thirty men; with peace
they could herd their livestock in distant places where forage was
good without fear of molestation. He proposed, in place of war, a
peaceful settlement of difficulties and trade with the Mormons.
After several days of consultation, peace was agreed upon. The
council appointed one of the chiefs, Hastele, who lived near the
Colorado River, as negotiator who ended by saying, “We hope we
may be able to eat at one table, be warmed by one fire, smoke one
pipe and sleep under one blanket.” Thus was peace promised, though
it was soon again to be put in jeopardy. Hamblin reached Kanab with
the good news about December 11, 1870.
[78]
Within a few weeks, a group of eighty Navajos arrived at Kanab on a
trading expedition. They came on foot and brought all the Navajo
blankets they could carry. They scattered among the settlements and
traded their blankets for horses and returned well satisfied with the
experiment.
Peaceful trading continued until the winter of 1873-74, when a party
of four young Navajos was caught in a snowstorm near a ranch in
Grass Valley, Sevier County. They made themselves at home at the
ranch and even killed a small animal for food. The owner of the
ranch, said to be a non-Mormon, learned of their presence and
gathered some of his friends to go with him to investigate. At the
ranch, they shot and killed three of the Indians and wounded
the fourth, who escaped and after painful hardships made his
way home.
[79]
His story inflamed Navajo vengefulness and disquieting reports
reached the Mormons of threatened reprisals. Brigham Young asked
Hamblin to visit the Navajos again and satisfy them that the Mormons
were not involved in the outrage. Bishop Levi Stewart of Kanab,
however, tried to dissuade Hamblin and even sent a messenger to
induce him to return after he had started, urging that the risk was
too great.
Firm in his purpose, Hamblin went his way and met the Navajos east
of Moencopi, about January 29, 1874. Hastele, the representative
appointed by the Navajos, was not there, but other influential Indians
considered Hamblin’s statement. The war council was held in a
Navajo hogan, to which there was but one entrance opposite Hamblin
and his two companions, while two dozen Navajos occupied the
space between.
Hamblin’s explanation of the killings was at first rejected on the
ground that it was he who had invited the Navajos to come into the
Mormon country to trade, with the result that three of their good
young men now lay on the ground “for the wolves to eat.” The
interpreter told Hamblin his companions could go home, but he must
die. The moment was tense. His companions refused to leave him.
Without arousing suspicion, Hamblin passed several revolvers to his
friends, saying as he did so, “These are in my way.” The men behind
unobtrusively readied them in case of emergency. Hamblin reminded
the Indians of his many friendly acts, of his willingness to come into
their midst to settle the matter, and told them it was not right to kill
him for the acts of strangers for whom he was not responsible. The
wounded Indian was brought in. A stirring appeal for revenge was
made by a young warrior, who demanded that Hamblin be the victim.
The Indians, however, after the excitement subsided, offered to settle
for three hundred and fifty horses and cattle. Hamblin deliberately
refused. One of them remarked that he would agree after he had
been stretched over the hot coals of the fire. The interpreter asked if
he were not afraid. “No,” he said, “my heart has never known fear.
What is there to scare me?” “The Navajos,” was their answer, to
which he replied that he “was not afraid of his friends.” Mollified, the
Indians finally agreed to leave the matter to be settled by Hastele
after an investigation.
Late that spring, Hastele and his party visited Kanab and were piloted
to Sevier Valley where their findings convinced them that the
Mormons were innocent. Thus ended the last threat to peaceful
179
relations with the Navajos. Thereafter, both groups traded on good
terms largely due to the outstanding bravery and cool judgment of
Jacob Hamblin.
Expansion in Kane County
Re-settlement of Long Valley and Kanab does not seem to have been
attempted until 1870, although Kanab and Paria were occupied by
missionaries under Jacob Hamblin in 1867 as frontier outposts. At
Paria a strong guard house and corral was built and some land was
cultivated, beginnings out of which the settlement grew with the
accession of several families in 1872 and 1873.
Kanab was similarly restored. The necessity of a fort there was
impressed upon the whites by the continued Navajo raids. Five stone
masons were sent from St. George in 1869 to construct the fort. They
reached Kanab on August 28 and worked until early in September,
when John R. Young told them they had finished their mission and
could go home.
This building expedition brought new settlers to Kanab, for John
Mangum (or Mangram), his brother, James, James Wilkins, and
George Ross, moved there soon after. Nate Adams, who visited
Kanab in September, 1870, and who moved there March 14, 1871,
says the first three were in hiding and that John D. Lee, also in
hiding, took up Scutumpah Ranch and explored Lee’s Ferry in 1869.
Several missionaries were sent to aid Hamblin about the same time.
They were fencing and cultivating land when Brigham Young made
his first visit to the Kanab country about the 1st of April, 1870.
George Albert Smith wrote of this visit:
At Kanab we met Brothers Jacob Hamblin and Jehiel MacConnel
[McConnell], and several other missionaries, who were engaged in
teaching the Indians how to cultivate the soil and to obtain a living
by peaceful pursuits. We were much pleased with the country.... As
soon as measures shall be taken to prevent the annual raids of the
180
Navajos, this land of Canaan will be re-occupied by the Saints and
become a valuable acquisition to our southern settlements.
[80]
A pioneer Salt Lake photographer, C. R. Savage, took many pictures
along the way, including one of Brigham Young and his party on the
Colorado at the mouth of the Virgin, and several of Zion Canyon.
Upon his return to Salt Lake City on April 16, 1870, Brigham Young
sent a group of fifty-two people led by Levi Stewart, to re-settle the
Kanab country. They went down through the Dixie settlements
and reached Pipe Springs on June 1. They remained there and
at Moccasin Springs several days while exploring the region.
[81]
On
the 14th they moved over to Kanab Creek and joined Jacob Hamblin
at the old fort, now too small to house so many.
Brigham Young manifested much interest in the success of the
colony. He promised Stewart that he would visit him in the fall and
asked him to find a more direct route to Kanab from the north that
would obviate the long roundabout approach through the Dixie
settlements and the Arizona strip. Stewart sent out two expeditions,
the second of which found a road from the head of the Sevier River
through Upper Kanab and Johnson Wash.
Brigham Young started for Kanab on August 26, 1870. Accompanying
him from Parowan was the intrepid explorer and topographer of the
U. S. Geological Survey, Major J. W. Powell, who had already made
one trip through the Grand Canyon and was returning to make plans
for further geological studies and his second trip through the canyon.
[82]
In attempting to follow Stewart’s directions, the party lost its way
and wandered into the Paria River valley and thus went many miles
out of its direct route. According to Nate Adams,
[83]
“old Humpy
Indian” guided the company safely into Kanab on the evening of
September 9, 1870. While there, a townsite and fields were surveyed
east of the fort beyond the path of the canyon winds. Brigham Young
returned to Salt Lake City via St. George and the Dixie settlements.
Three months later, on December 14, 1870, six lives were lost in a
fire at the fort. These included Bishop Stewart’s wife, Margery, and
three of his sons.
[84]
Brigham Young made a special trip to Kanab
from St. George, where he was wintering, to comfort the bereaved
families. Soon after, the settlers began to build their homes on the
townsite. Within a few years, the fort was deserted but it was
maintained for some time for use in case of emergency. Dellenbaugh,
a member of Major J. W. Powell’s party, thus describes his visit to
Kanab in the early 70’s:
F. S. Dellenbaugh in Zion Canyon (1930) with the flag of the
Emma Dean boat that made “a canyon voyage” about 60 years
Photo U. S. Nat. Park Service.
Courtesy Union Pacific Railroad.
181
earlier.
Viewing Bryce Canyon from the rim.
... Nigger, [a white mule] went along very well and I was in
Kanab by three o’clock. The village which had been started
only a year or two, was laid out in the characteristic Mormon style,
with wide streets and regular lots fenced by wattling willows
between stakes. Irrigating ditches ran down each side of every
street and from them the water, derived from a creek that came
down a canyon back of the town, could be led into any of the lots,
each of which was about one quarter of an acre; that is, there
were four lots to a block. Fruit trees and vines had been planted
and were already beginning to promise near results, while corn,
182
potatoes, etc., gave fine crops. The original place of settlement was
a square formed by one-story log houses on three sides and a
stockade on the fourth. This was called the fort and was a place of
refuge though the danger from Navajo attack seemed to be over
and that from any assault by the Paiutes certainly was past. One
corner of the fort was made by the walls of the schoolhouse, which
was at the same time meeting-house and ball-room. Altogether
there were about 100 families in the village. The houses that had
been built outside the fort were quite substantially constructed,
some of adobe or sun-dried brick. The entire settlement had a
thrifty air, as is the case with the Mormons. Not a grog-shop, or
gambling saloon, or dance-hall was to be seen; quite in contrast
with the usual disgraceful accompaniments of the ordinary frontier
towns. A perfectly orderly government existed, headed by a bishop
appointed by the church authorities in Salt Lake City, the then
incumbent of this office being an excellent man, Bishop Stewart.
[85]
After the Navajo peace settlement many of the places abandoned in
1866 were reoccupied and within a few years further expansion filled
most of the remaining areas suitable for settlements or ranches. In
Long Valley, Berryville (now Glendale), and Winsor (Mt. Carmel) were
revived in 1871. Johnson was settled in the spring of the same year
by five brothers, Joel, Benjamin, Joseph, George and William
Johnson, on the site of Scutumpah, formerly John D. Lee’s ranch. In
1872, Graham, on the headwaters of Kanab Creek (upper Kanab),
was reoccupied and the settlers engaged in dairying and lumbering.
The upper reaches of the Paria, however, attracted settlers from the
north. Panguitch was re-founded in March, 1871 under George W.
Sevy and counted seventy-five families the next year. Joel H. Johnson
and George D. Wilson established a sawmill in 1871 near the present
location of Hillsdale, and were soon joined by twenty families,
including those of Nephi and Seth Johnson. Other cattlemen located
farther up the Sevier, where Meltiar Hatch founded the village
bearing his name. Nephi Johnson, discoverer of Zion Canyon,
was made bishop of Hillsdale in 1874.
Attention was then focused on the upper Paria. The first settlers,
David O. Littlefield and Orley D. Bliss, located near the present site of
Cannonville on Christmas Eve, 1874. Early the next day eight other
families arrived, who built log houses at a place called Clifton and
began farming along the Paria and on Henrieville Creek. Ebenezer
Bryce, from Pine Valley, selected a place farther upstream, a mile or
two east of the present site of Tropic near the mouth of Bryce
Canyon. Bryce used the famous canyon for a cattle range, and thus
immortalized his name.
Clifton was not well located and in 1877 some settlers moved to a
new townsite called Cannonville, in honor of George Q. Cannon, high
Mormon official who had taken a special interest in their affairs.
Other settlers moved over to Henrieville Creek to be near their farms,
and thus the town of Henrieville (named for James Henrie, president
of Panguitch Stake) was born.
In 1879 Daniel Goulding settled near Bryce’s ranch. Seeking water for
irrigation, he and Bryce devised a scheme to divert water from Pine
Creek in the Great Basin by means of a canal over the divide. This
they finished, but upkeep was expensive, their crops were poor, and
Goulding lost about five hundred fruit trees from drouth. Bryce
became discouraged and left for Arizona in 1880 and Goulding moved
to Henrieville in 1883. Bryce, unimpressed by the beauty of the
canyon, considered it “awful hard to find a cow that was lost” in the
intricate maze of its pinnacles.
Seth Johnson and several others located in 1886 on Yellow Creek
(Kane County) about three miles southwest of Cannonville and
named the settlement Georgetown, in honor of the same man for
whom Cannonville was named.
In 1890 the two Ahlstrom brothers built homes on the present site of
Tropic and with several others began a second and more ambitious
attempt to divert water from the East Fork of Sevier River over the
divide into Paria Creek. This time the project succeeded. Tropic
townsite was surveyed in 1891 and settlers began to flock there and
183
prepare homes and lands in anticipation of the coming of the water. A
fitting celebration was staged on May 23, 1892, when the water was
turned into the canal.
By this time, most of the suitable valleys and canyons had been
occupied. Erosion, however, caused trouble at Kanab. From 1883 to
1890, floods presumably resulting from overgrazing tore out dams
and ditches and gutted the canyons and valleys with deep washes.
Water arose in the bottom of the washes and that in Kanab Wash
(below Kanab) was diverted about 1886 onto a new townsite just
beyond the state line in Arizona, called Fredonia, which later served
as a refuge for a number of polygamous wives during the
Federal offensive against the practice.
[86]
While southern Utah was thus growing, a new movement was
developed. In 1879, the Mormon Church leaders called for eighty
men from the Southern Mission to establish an outpost for the
purpose of “cultivating and maintaining friendly relations with Indians
whose homes were near the point where the state of Colorado and
the Territories of Utah, New Mexico and Arizona come together.”
Twenty-five men, including Kumen Jones, went out to investigate
routes and locations. They traveled via Lee’s Ferry, Tuba City and
Monument Valley to the San Juan River as far as Four-Corners, spent
about three months exploring the region, and then returned home via
a northern route, past the sites of Monticello and Moab.
[87]
While they were away, another party set out from Escalante seeking
a short-cut to the San Juan country. A route much more direct than
that mapped by the first party was reported, apparently on imperfect
observation. It was, however, accepted, and by October the party
was on its way. It passed through Escalante and reached Forty-mile
Spring where it was held up by excessively rough country, while
snows in the mountains blocked retreat.
Three scouts were sent ahead to investigate some of the wildest and
most rugged scenic areas of America. The three returned in
184
disappointment; one held the route feasible, another positively
rejected it, while the third thought it might be possible to get through
with special help. Envoys were sent to Salt Lake City to appeal for
assistance, which was given in the form of a legislative appropriation
for blasting a way through.
It took fifty days to get eighty-two wagons through Hole-in-the-Rock
and down to the Colorado River and ninety days to reach Bluff on the
San Juan River where the first settlement was made. Three babies
were born on the way and the hardships endured form a Western
epic.
[88]
The story of Orderville has been left for the last. The United Order
[89]
was organized at Mt. Carmel, March 20, 1874, by John R.
Young, at which time one hundred and nine members were
listed. One summer of the United Order was enough for most
members. Bishop Bryant Jolley, with his numerous family and
relatives, formed the core of the dissenters. To avoid contention,
those who wished to continue with the Order sold their holdings and
moved in a body two miles above Mt. Carmel where title to all land
was vested in the group and where they set up the town Orderville,
under the leadership of Howard O. Spencer. The new town was
surveyed February 20, 1875.
The first building was a hotel where all ate together in the large
dining hall, from July, 1875 to May, 1880. As time passed, living
quarters were provided for each family and work was divided into
more specialized fields.
During the hey-day of the Order, around 1880, it numbered nearly six
hundred adherents and there were some twenty-eight specialized
departments of work which included most of the various activities
that go to make up a simple community. The Order made great
progress and acquired property rapidly. Farming lands were expanded
to include areas scattered through Long Valley and at Kanab.
185
The growing power of the Order created jealousies, but disintegration
came from internal dissension. The idea of giving everyone an equal
reward regardless of ability or accomplishment tended in many cases
to lessen effort and brought charges of laziness and carelessness.
Gradually, more and more individual property was assigned to each
home; farmers were given a share of their own produce and livestock
and sawmills and freighting were operated under lease or contract.
Matters were hastening toward dissolution when, in 1885, polygamy
troubles began. Fear that the Federal government might confiscate
the goods of the Order forced the final dissolution of most of the
property, and farming lands, livestock, ranches, tannery and sawmill,
were all sold to members. The woolen mill alone was kept and
intermittently operated until 1900. In that year the United Order of
Orderville was officially dissolved, twenty-five years after its
incorporation.
Zion Canyon
“In an instant, there flashed before us a scene never to be forgotten.
In coming time it will, I believe, take rank with a very small number
of spectacles each of which will, in its own way, be regarded as the
most exquisite of its kind which the world discloses. The scene before
us was the Temples and Towers of the Virgin.” Thus prophetically
wrote Captain Clarence E. Dutton of the U. S. Geological Survey in a
report published in the year 1880.
Dutton was following up the geological work begun by Major J. W.
Powell ten or twelve years earlier when the latter started out to
explore the Colorado River and made his two memorable trips in
boats down the river through the Grand Canyon. The geological
problems encountered were so extraordinary that Dutton was
detailed to further investigation and encountered problems that have
engaged the attention of scientists to this day, particularly the
eminent geologist, Dr. Herbert E. Gregory.
Following the line of Vermillion Cliffs from Kanab westward, Dutton
came to the pass between Eagle Crags and Smithsonian Butte when
suddenly, startlingly, there lay before him to the northward the valley
of the Virgin River in all its grandeur.
Few have seen Zion as Clarence Dutton saw it. From a high pass, in
late afternoon, with the sun on his left, he looked into that vast
panorama of the Vermillion Cliffs of Zion and Parunuweap and those
flanking the Great West Canyon as well—a twenty-mile stretch in one
sweeping view. The setting sun cast shadows that made the turrets
and towers stand out in bold relief, while the light reflected from one
wall upon another intensified the tints and shades of the reds until
they stood out in striking contrast with the vivid green of the
186
vegetation and the higher cliffs. No wonder the cold scientist broke
down and described in emotional terms this superb panorama.
Forerunners of Dutton had visited Zion, but none had penned such
eloquent praise. Major J. W. Powell and two companions, Stephen V.
Jones, one of his topographers, and Joseph W. Young, a Mormon, left
Long Valley on September 10, 1872
[90]
and started down through the
Parunuweap on foot. They came out next day before noon and spent
another day visiting Zion Canyon. Of this trip, Powell says of the
Parunuweap:
At noon, we are in a canyon 2500 feet deep and we come to a fall
where the walls are broken down, and the huge rocks beset the
channel, on which we obtain a foothold to reach a level two
hundred feet below. Here the canyon is again wider and we
find a floodplain on which we can walk.
Next day of the Mukuntuweap Canyon, he writes:
Entering this, we have to wade up the stream; often the water fills
the entire channel, and although we travel many miles, we find no
floodplain, talus, or broken piles of rock at the foot of the cliff. The
walls have smooth, plain faces, and are everywhere very regular
and vertical for a thousand feet or more, where they seem to break
in shelving slopes to higher altitudes; and everywhere as we go
along, we find springs bursting out at the foot of the walls.
[91]
Jack Hillers, a photographer from Powell’s party, spent some time
during the summer of 1873 taking pictures in Zion Canyon. These are
on file in the U. S. Geological Survey Office and have been often used
in publications. For a long time, however, this material and Zion
Canyon were largely forgotten. The local course of development
continued placidly for many years. Only occasionally a hardy traveler,
hearing of the beauties of the region, had courage enough to brave
the rocky, dusty roads to enjoy the scenic splendors. One was
Frederick S. Dellenbaugh, who had accompanied Powell on his
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The Zebrafish Genetics Genomics and Transcriptomics 4th Edition H. William Detrich

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  • 2. We have selected some products that you may be interested in Click the link to download now or visit ebookultra.com for more options!. The Zebrafish Genetics Genomics and Informatics 3rd Edition H. William Detrich https://ebookultra.com/download/the-zebrafish-genetics-genomics-and- informatics-3rd-edition-h-william-detrich/ The Zebrafish Disease Models and Chemical Screens 3rd Edition H. William Detrich https://ebookultra.com/download/the-zebrafish-disease-models-and- chemical-screens-3rd-edition-h-william-detrich/ Cardiovascular Genetics and Genomics 1st Edition Dan M. Roden https://ebookultra.com/download/cardiovascular-genetics-and- genomics-1st-edition-dan-m-roden/ Encyclopedia of Genetics Genomics Proteomics and Bioinformatics 1st Edition Michael J. Dunn https://ebookultra.com/download/encyclopedia-of-genetics-genomics- proteomics-and-bioinformatics-1st-edition-michael-j-dunn/
  • 3. Quantitative Genetics Genomics and Plant Breeding Cabi First Edition M S Kang https://ebookultra.com/download/quantitative-genetics-genomics-and- plant-breeding-cabi-first-edition-m-s-kang/ Genetics Genomics and Breeding of Cucurbits 1st Edition Yi-Hong Wang (Editor) https://ebookultra.com/download/genetics-genomics-and-breeding-of- cucurbits-1st-edition-yi-hong-wang-editor/ Genetics Genomics and Breeding of Grapes 1st Edition Anne- Francoise Adam-Blondon https://ebookultra.com/download/genetics-genomics-and-breeding-of- grapes-1st-edition-anne-francoise-adam-blondon/ Flax Lipids Classes Biosynthesis Genetics and the Promise of Applied Genomics for Understanding and Altering of Fatty Acids Classes Biosynthesis Genetics and the Promise of Applied Genomics for Understanding and Altering of Fatty Acids 1st Edition Bourlaye Fofana https://ebookultra.com/download/flax-lipids-classes-biosynthesis- genetics-and-the-promise-of-applied-genomics-for-understanding-and- altering-of-fatty-acids-classes-biosynthesis-genetics-and-the-promise- of-applied-genomics-for-unders/ Emery and Rimoin s Principles and Practice of Medical Genetics and Genomics Foundations Reed E Pyeritz https://ebookultra.com/download/emery-and-rimoin-s-principles-and- practice-of-medical-genetics-and-genomics-foundations-reed-e-pyeritz/
  • 5. The Zebrafish Genetics Genomics and Transcriptomics 4th Edition H. William Detrich Digital Instant Download Author(s): H. WilliamDetrich, Monte Westerfield and Leonard I. Zon (Eds.) ISBN(s): 9780128034743, 0128034742 Edition: 4 File Details: PDF, 20.70 MB Year: 2016 Language: english
  • 6. Methods in Cell Biology The Zebrafish: Genetics, Genomics, and Transcriptomics Volume 135
  • 7. Series Editors Leslie Wilson Department of Molecular, Cellular and Developmental Biology University of California Santa Barbara, California Phong Tran University of Pennsylvania Philadelphia, USA & Institut Curie, Paris, France
  • 8. Methods in Cell Biology The Zebrafish: Genetics, Genomics, and Transcriptomics Volume 135 Edited by H. William Detrich, III Northeastern University Marine Science Center, Nahant, MA, United States Monte Westerfield University of Oregon, Eugene, OR, United States Leonard I. Zon Harvard University, Boston, MA, United States AMSTERDAM • BOSTON • HEIDELBERG • LONDON NEW YORK • OXFORD • PARIS • SAN DIEGO SAN FRANCISCO • SINGAPORE • SYDNEY • TOKYO Academic Press is an imprint of Elsevier
  • 9. Academic Press is an imprint of Elsevier 50 Hampshire Street, 5th Floor, Cambridge, MA 02139, USA 525 B Street, Suite 1800, San Diego, CA 92101-4495, USA 125 London Wall, London EC2Y 5AS, UK The Boulevard, Langford Lane, Kidlington, Oxford OX5 1GB, UK Fourth edition 2016 Copyright © 2016, 2011, 2004, 1999 Elsevier Inc. All rights reserved. No part of this publication may be reproduced or transmitted in any form or by any means, electronic or mechanical, including photocopying, recording, or any information storage and retrieval system, without permission in writing from the publisher. Details on how to seek permission, further information about the Publisher’s permissions policies and our arrangements with organizations such as the Copyright Clearance Center and the Copyright Licensing Agency, can be found at our website: www.elsevier.com/permissions. This book and the individual contributions contained in it are protected under copyright by the Publisher (other than as may be noted herein). Notices Knowledge and best practice in this field are constantly changing. As new research and experience broaden our understanding, changes in research methods, professional practices, or medical treatment may become necessary. Practitioners and researchers must always rely on their own experience and knowledge in evaluating and using any information, methods, compounds, or experiments described herein. In using such information or methods they should be mindful of their own safety and the safety of others, including parties for whom they have a professional responsibility. To the fullest extent of the law, neither the Publisher nor the authors, contributors, or editors, assume any liability for any injury and/or damage to persons or property as a matter of products liability, negligence or otherwise, or from any use or operation of any methods, products, instructions, or ideas contained in the material herein. ISBN: 978-0-12-803474-3 ISSN: 0091-679X For information on all Academic Press publications visit our website at https://www.elsevier.com Publisher: Zoe Kruze Acquisition Editor: Zoe Kruze Editorial Project Manager: Sarah Lay Production Project Manager: Malathi Samayan Designer: Victoria Pearson Typeset by TNQ Books and Journals
  • 10. Len, Monte, and I dedicate the 4th Edition of Methods in Cell Biology: The Zebrafish to the postdoctoral fellows and graduate students who conducted the genetic screens that established the zebrafish as a preeminent vertebrate model system for analysis of development.
  • 11. Contributors J. Ablain Howard Hughes Medical Institute and Harvard Medical School, Boston, MA, United States K. Asakawa SOKENDAI (The Graduate University for Advanced Studies), Mishima, Shizuoka, Japan H. Ata Mayo Clinic, Rochester, MN, United States J. Bakkers Hubrecht Institute and University Medical Centre Utrecht, Utrecht, The Netherlands J. Bessa IBMC-Instituto de Biologia Molecular e Celular, Porto, Portugal; Universidade do Porto, Porto, Portugal Y.M. Bradford University of Oregon, Eugene, OR, United States B.R. Cairns University of Utah School of Medicine, Salt Lake City, UT, United States W. Chen Vanderbilt University School of Medicine, Nashville, TN, United States J. Cibelli Michigan State University, East Lansing, MI, United States; BIONAND, Andalucı́a, Spain K.J. Clark Mayo Clinic, Rochester, MN, United States P. Coucke Ghent University, Ghent, Belgium V.T. Cunliffe University of Sheffield, Sheffield, United Kingdom F. Del Bene PSL Research University, Paris, France xvii
  • 12. A. De Paepe Ghent University, Ghent, Belgium F. De Santis PSL Research University, Paris, France V. Di Donato PSL Research University, Paris, France A. Eagle University of Oregon, Eugene, OR, United States S.C. Ekker Mayo Clinic, Rochester, MN, United States T. Erickson Oregon Health & Science University, Portland, OR, United States T. Evans Weill Cornell Medical College, New York, NY, United States D. Fashena University of Oregon, Eugene, OR, United States A. Felker University of Zürich, Zürich, Switzerland A. Fernández-Miñán Centro Andaluz de Biologı́a del Desarrollo (CABD), Consejo Superior de Investigaciones Cientı́ficas/Universidad Pablo de Olavide/Junta de Andalucı́a, Sevilla, Spain K. Frazer University of Oregon, Eugene, OR, United States A. Ghosha Carnegie Institution for Science, Baltimore, MD, United States M.G. Goll Memorial Sloan Kettering Cancer Center, New York, NY, United States J.L. Gómez-Skarmeta Centro Andaluz de Biologı́a del Desarrollo (CABD), Consejo Superior de Investigaciones Cientı́ficas/Universidad Pablo de Olavide/Junta de Andalucı́a, Sevilla, Spain a Current address: Tata Institute of Fundamental Research, Mumbai, India xviii Contributors
  • 13. D.J. Grunwald University of Utah, Salt Lake City, UT, United States M.E. Halpern Carnegie Institution for Science, Baltimore, MD, United States J.K. Heath Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia K. Hoshijima University of Utah, Salt Lake City, UT, United States D.G. Howe University of Oregon, Eugene, OR, United States H. Huang University of California Los Angeles, Los Angeles, CA, United States J.P. Junker Hubrecht Institute and University Medical Centre Utrecht, Utrecht, The Netherlands M.J. Jurynec University of Utah, Salt Lake City, UT, United States P. Kalita University of Oregon, Eugene, OR, United States K. Kawakami SOKENDAI (The Graduate University for Advanced Studies), Mishima, Shizuoka, Japan M.C. Keightley Australian Regenerative Medicine Institute, Clayton, VIC, Australia; Monash University, Clayton, VIC, Australia F. Kruse Hubrecht Institute and University Medical Centre Utrecht, Utrecht, The Netherlands C. Lawrence Boston Children’s Hospital, Boston, MA, United States S. Lefever Ghent University, Ghent, Belgium C. Li Memorial Sloan Kettering Cancer Center, New York, NY, United States; Weill Cornell Graduate School of Medical Sciences, New York, NY, United States Contributors xix
  • 14. G.J. Lieschke Australian Regenerative Medicine Institute, Clayton, VIC, Australia; Monash University, Clayton, VIC, Australia S. Lin University of California Los Angeles, Los Angeles, CA, United States C.G. Love Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia; University of Melbourne, Parkville, VIC, Australia L.A. Maddison Vanderbilt University School of Medicine, Nashville, TN, United States P. Mani University of Oregon, Eugene, OR, United States S. Markmiller University of California San Diego, La Jolla, CA, United States R. Martin University of Oregon, Eugene, OR, United States S. Masuda Tokyo Institute of Technology, Yokohama, Japan A.C. Miller University of Oregon, Eugene, OR, United States C.B. Moens Fred Hutchinson Cancer Research Center, Seattle, WA, United States C. Mosimann University of Zürich, Zürich, Switzerland S.T. Moxon University of Oregon, Eugene, OR, United States M.C. Mullins University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, United States P.J. Murphy University of Utah School of Medicine, Salt Lake City, UT, United States K.N. Murray University of Oregon, Eugene, OR, United States xx Contributors
  • 15. A. Muto SOKENDAI (The Graduate University for Advanced Studies), Mishima, Shizuoka, Japan T. Nicolson Oregon Health & Science University, Portland, OR, United States C.J. Ott Dana Farber Cancer Institute, Harvard Stem Cell Institute, Boston, MA, United States H. Paddock University of Oregon, Eugene, OR, United States F. Pelegri University of WisconsineMadison, Madison, WI, United States C. Pich University of Oregon, Eugene, OR, United States S. Prukudom Kasetsart University, Bangkok, Thailand; Center of Excellence on Agricultural Biotechnology: (AG-BIO/PERDO-CHE), Bangkok, Thailand S. Ramachandran University of Oregon, Eugene, OR, United States J.E.J. Rasko Centenary Institute, Camperdown, NSW, Australia; University of Sydney, Sydney, NSW, Australia; Royal Prince Alfred Hospital, Newtown, NSW, Australia M.P. Rossmann Harvard University, Harvard, Cambridge, MA, United States L. Ruzicka University of Oregon, Eugene, OR, United States K. Schaper University of Oregon, Eugene, OR, United States A.N. Shah Fred Hutchinson Cancer Research Center, Seattle, WA, United States X. Shao University of Oregon, Eugene, OR, United States A. Singer University of Oregon, Eugene, OR, United States Contributors xxi
  • 16. K. Siripattarapravat Kasetsart University, Bangkok, Thailand; Center of Excellence on Agricultural Biotechnology: (AG-BIO/PERDO-CHE), Bangkok, Thailand F. Speleman Ghent University, Ghent, Belgium M. Superdock Boston Children’s Hospital, Boston, MA, United States; Dana Farber Cancer Institute, Harvard Stem Cell Institute, Boston, MA, United States; Harvard Medical School and Howard Hughes Medical Institute, Boston, MA, United States M. Tanaka Tokyo Institute of Technology, Yokohama, Japan J.J. Tena Centro Andaluz de Biologı́a del Desarrollo (CABD), Consejo Superior de Investigaciones Cientı́ficas/Universidad Pablo de Olavide/Junta de Andalucı́a, Sevilla, Spain S. Toro University of Oregon, Eugene, OR, United States A. van Oudenaarden Hubrecht Institute and University Medical Centre Utrecht, Utrecht, The Netherlands J. Vandesompele Ghent University, Ghent, Belgium S. Vanhauwaert Ghent University, Ghent, Belgium C. Van Slyke University of Oregon, Eugene, OR, United States Z.M. Varga University of Oregon, Eugene, OR, United States H. Wada SOKENDAI (The Graduate University for Advanced Studies), Mishima, Shizuoka, Japan M. Westerfield University of Oregon, Eugene, OR, United States xxii Contributors
  • 17. A. Willaert Ghent University, Ghent, Belgium S. Yang Boston Children’s Hospital, Boston, MA, United States; Dana Farber Cancer Institute, Harvard Stem Cell Institute, Boston, MA, United States; Harvard Medical School and Howard Hughes Medical Institute, Boston, MA, United States L. Yin Vanderbilt University School of Medicine, Nashville, TN, United States B. Zhang Peking University, Beijing, People’s Republic of China Y. Zhang Peking University Shenzhen Graduate School, Shenzhen, China; University of California Los Angeles, Los Angeles, CA, United States Y. Zhou Boston Children’s Hospital, Boston, MA, United States; Dana Farber Cancer Institute, Harvard Stem Cell Institute, Boston, MA, United States; Harvard Medical School and Howard Hughes Medical Institute, Boston, MA, United States; Harvard University, Harvard, Cambridge, MA, United States L.I. Zon Boston Children’s Hospital, Boston, MA, United States; Dana Farber Cancer Institute, Harvard Stem Cell Institute, Boston, MA, United States; Harvard Medical School and Howard Hughes Medical Institute, Boston, MA, United States; Harvard University, Harvard, Cambridge, MA, United States Contributors xxiii
  • 18. Preface Len, Monte, and I are pleased to introduce the fourth edition of Methods in Cell Biology: The Zebrafish. The advantages of the zebrafish, Danio rerio, are numerous, including its short generation time and high fecundity, external fertilization, and the optical transparency of the embryo. The ease of conducting forward genetic screens in the zebrafish, based on the pioneering work of George Streisinger, culminated in screens from the laboratories of Wolfgang Driever, Mark C. Fishman, and Christiane Nüsslein-Volhard, published in a seminal volume of Development (volume 123, December 1, 1996) that described a “candy store” of mutants whose phenotypes spanned the gamut of developmental processes and mechanisms. Life for geneticists who study vertebrate development became really fine. Statistics derived from ZFIN (the Zebrafish Model Organism Database; http://zfin. org) illustrate the dramatic growth of research involving zebrafish. The zebrafish genome has been sequenced, and as of 2014, more than 25,000 genes have been placed on the assembly. Greater than 15,500 of these genes have been established as orthologs of human genes. The zebrafish community has grown from w1,400 re- searchers in 190 laboratories as of 1998 to w7,000 in 930 laboratories in 2014. The annual number of publications based on the zebrafish has risen from 1,913 to 21,995 in the same timeframe. Clearly, the zebrafish has arrived as a vertebrate biomedical model system par excellence. When we published the first edition (volumes 59 and 60) in 1998, our goal was to encourage biologists to adopt the zebrafish as a genetically tractable model organism for studying biological phenomena from the cellular through the organismal. Our goal today remains unchanged, but the range of subjects and the suite of methods have expanded rapidly and significantly in sophistication over the years. With the second and third editions of MCB: The Zebrafish (volumes 76 and 77 in 2004; volumes 100, 101, 104, and 105 in 2010e11), we documented this extraordinary growth, again relying on the excellent chapters contributed by our generous col- leagues in the zebrafish research community. When Len, Monte, and I began planning the fourth edition, we found that the zebrafish community had once more developed and refined novel experimental systems and technologies to tackle challenging biological problems across the spectrum of the biosciences. We present these methods following the organizational structure of the third edition, with volumes devoted to Cellular and Developmental Biology, to Genetics, Genomics, and Transcriptomics, and to Disease Models and Chemical Screens. Here we introduce the third volume, Genetics, Genomics, and Transcriptomics. Genetics, Genomics, and Transcriptomics is divided into five sections that cover genetic and genomics techniques. Part 1 covers forward and reverse genetics in nine chapters, many of which employ the revolutionary CRISPR/Cas9 technology in novel ways. Precision editing of the zebrafish genome through homologous recom- bination has now become a reality. Part 2 contains five chapters that describe xxv
  • 19. advances in transgenesis and functional genomics approaches. Spatially resolved transcriptomics at the organismal level, cell type-specific transcriptomics, and the important companion technology, RT-qPCR, are presented in Part 3. We devote Part 4 to five chapters on the emerging analysis of epigenetic regulation of gene expression in the zebrafish. Part 5 concludes the volume with four important chap- ters on zebrafish husbandry, health monitoring, disease prevention, and information technology. We anticipate that you, our readership, will apply these methods successfully in your own zebrafish research programs and will develop your own technical advances that may be considered for a future edition of Methods in Cell Biology: The Zebra- fish. The zebrafish is a remarkable experimental systemdthe preeminent vertebrate model for mechanistic studies of cellular and developmental processes in vivo. We thank the series editors, Leslie Wilson and Phong Tran, and the staff of Elsevier/Academic Press, especially Zoe Kruze and Sarah Lay, for their enthusiastic support of our fourth edition. Their help, patience, and encouragement are pro- foundly appreciated. H. William Detrich, III Monte Westerfield Leonard I. Zon xxvi Preface
  • 20. Multiplex conditional mutagenesis in zebrafish using the CRISPR/Cas system 1 L. Yina , L.A. Maddisona , W. Chen1 Vanderbilt University School of Medicine, Nashville, TN, United States 1 Corresponding author: E-mail: wenbiao.chen@vanderbilt.edu CHAPTER OUTLINE Introduction................................................................................................................ 4 1. Methods ................................................................................................................ 5 1.1 Assembly of U6-Based sgRNA Transgenic Constructs................................. 5 1.2 Construction of Cas9 Expression Vectors ................................................... 9 1.3 Screening and Evaluation of Stable sgRNA or Cas9 Transgenic Fish ............ 9 2. Discussion........................................................................................................... 14 Summary .................................................................................................................. 14 Acknowledgments..................................................................................................... 15 References ............................................................................................................... 15 Abstract The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-asso- ciated protein (Cas) system is a powerful tool for genome editing in numerous organisms. However, the system is typically used for gene editing throughout the entire organism. Tissue and temporal specific mutagenesis is often desirable to determine gene function in a specific stage or tissue and to bypass undesired consequences of global mutations. We have developed the CRISPR/Cas system for conditional mutagenesis in transgenic zebrafish using tissue-specific and/or inducible expression of Cas9 and U6-driven expression of sgRNA. To allow mutagenesis of multiple targets, we have isolated four distinct U6 promoters and designed Golden Gate vectors to easily assemble transgenes with multiple sgRNAs. We provide experimental details on the reagents and applications for multiplex conditional mutagenesis in zebrafish. CHAPTER a These authors contributed equally. Methods in Cell Biology, Volume 135, ISSN 0091-679X, http://dx.doi.org/10.1016/bs.mcb.2016.04.018 © 2016 Elsevier Inc. All rights reserved. 3
  • 21. Exploring the Variety of Random Documents with Different Content
  • 22. 169 charge and brought with him forty-seven men under James Andrus with wagons and supplies for an extended trip designed to drive the Navajos across the Colorado River. When they arrived at Pipe Springs, the snow was two feet deep and no trace of the sheep or men could be found. On January 18, they came upon the tracks of two Paiute Indians following a large steer, tracked them until sundown, and captured the Indians in the act of killing the beef. After questioning and torture, hanging by the heels and twisting of thumbs, one of the Indians admitted that he had dreamed that Navajos had been there and then revealed the whereabouts of a camp of Indians about ten miles out. A detachment was sent and found that it had been moved another five miles. The militia overtook the camp about sunrise on January 20, killing two Indians and capturing five. Third degree methods elicited information about the killing of Whitmore and McIntyre. The captives led another detachment to the scene of the killings, where the posse crisscrossed the area on horseback, uncovering the arm of one of the victims in the deep snow. Both bodies had bullet wounds and were riddled with arrows. They had been killed on January 10. A wagon was sent after the bodies. While the men were recovering the remains the other detachment with the five Indian prisoners arrived. These had in their possession much of the clothing and personal effects of the murdered men. The evidence of guilt seemed conclusive, so the Indians were turned loose and shot as they attempted to run. The Navajos who probably assisted in the killing escaped. The sheep could not be found and it was assumed the Navajos had taken them across the Colorado River. As pursuit was impossible because of the deep snow the party returned home. Charles L. Walker of St. George records in his diary: They were brought home in a wagon load of snow, frozen stiff and in a good state of preservation. I, with others, washed them and pulled out the arrow points from their bodies and dressed them in
  • 23. their burial robes. Also went to the funeral, which was attended by a large concourse of people. [70] On February 19, 1866, two days after Erastus Snow was elected Brigadier General, Peter Shurtz, who had built a station at Paria and had kept about twenty Indians around him all winter, reported that he had lost his cattle and wished to move into the settlements. He also reported Navajos camped on Paria River about eight miles below his ranch where the Ute trail reached the stream. Further information indicated that the Navajos were concentrated east of the Colorado at Cottonwood, intending to raid Kane County in force and that Captain James Andrus with thirty men had gone to Paria to get Peter Shurtz and his family and to reconnoiter. No report of this expedition is available, but a letter written by L. W. Roundy from Kanab on March 9, 1866, tells that Andrus had left Paria fourteen days earlier headed for an Indian camp twelve miles south. [71] At Kanab, three Indians had attempted to kill Oren Clark in the bottoms near the fort and had started to drive off the livestock. Four men from the fort rushed in pursuit and after dark recovered about thirty head of cattle, but the Indians escaped with about an equal number. The Indian menace was so serious by this time that Erastus Snow ordered all stock in the region south of St. George and the Virgin River as far east as Kanab removed to the north and west of the lines of settlements so that it would be easier to ward off Navajo attacks. This was a difficult task because the grazing was poor around the settlements and the mountains to the northwest were already filled with livestock. The threat from the Utes in upper Sevier Valley also became acute. Menacing behavior of the Indians in this area and in the Kanab region led to an order from Utah headquarters to General Erastus Snow (March 15) to send a company of men from Beaver and Iron counties over to the Sevier River to build and man an outpost between
  • 24. 170 Circleville and Panguitch. A company of 76 men led by Captain Silas S. Smith served here from March 21 to November 30, 1866. They established Fort Sanford about ten miles north of Panguitch and assisted settlers at Circleville to move to safety. At Panguitch, they helped the settlers transform the town into a fort. In the meantime, gathering the livestock from the exposed range was proceeding slowly. A party sent out from Rockville in April to round up the stock in the vicinity of Maxwell’s Ranch, found the bodies of two men, a woman, and an Indian, killed a few days before. When the bodies were brought in to Grafton, it was ascertained that they were young Robert Berry, his wife, Isabel, and his brother, Joe, who were coming home to Berryville in Long Valley via the Dixie settlements and the Arizona Strip (a roundabout way, but the only wagon route at the time). They had left the Maxwell Ranch on Short Creek on April 2, 1866 when some Indians (presumably Paiutes), ambushed them. According to verbal reports, as related by Mrs. John Dennett of Rockville (then a girl living in Long Valley and who pieced her story from Indian and white sources), the Berrys fought for their lives. The Indians shot one of the horses, rendering the wagon useless. In the fighting, one Indian was shot. Joe Berry loosened the other horse and tried to escape but was killed in so doing. The Indians closed in and captured Robert and his wife. They tied Robert to a wheel where he was forced to watch them torture Isabel, who was an expectant mother. They shot arrows into her and laughed at her as she tried to pull them out. Then they shot him full of arrows. Mrs. Dennett said her father always felt that the Berrys had been killed in revenge for some Indians slain by Long Valley men who had found them roasting a beef. At that time three were slain: an Indian, a squaw and a papoose. When the Berry tragedy was reported in St. George, orders were issued forbidding travel unless in groups large enough to provide adequate safeguards. This led to the declaration of martial law, May 2, 1866, and to the issuance of instructions to concentrate the
  • 25. 171 settlers in fortified places of at least 150 men. Patrols were ordered out in various directions, especially across the trails used by the Navajos in raiding the Mormon country and in contacting the rampaging Utes of Sevier County. When Silas S. Smith, stationed on the Sevier, heard of the Berry massacre, he found that the Paiute chief at Panguitch had known about it for five days without reporting it to him. Smith at once ordered pickets to bring in all passing Indians for questioning. Friendly Indians responded willingly enough, but when two strange Indians refused, a skirmish resulted in which one was killed and the other wounded. Smith decided to disarm the local Indians and surrounded one of their camps near Panguitch one morning before daylight and took their arms. Two visiting Indians were missing from the camp so he kept a guard awaiting their arrival. When they came, they showed fight. One of them was killed, whereupon the other surrendered. The next day Smith surrounded another camp soon after sunrise, but the natives had already fled. However, in the ensuing melee two more Indians were killed. The arms taken from them included several guns, many new arrows, and a peck of new arrow heads. Some escaped to Panguitch Lake and spread the alarm among the Indians there. General Snow had a number of chiefs from Panguitch, Parowan, and Red Creek brought to Parowan for conference. He tried to pacify them with arguments and presents but insisted that they must not have arms or ammunition and must have passes in order to travel through the Mormon settlements. This aroused some resentment, but on April 25, 1866, they agreed to leave their weapons at Parowan as a token of friendship. Some of the Indians reported gunfire around Upper Kanab where Col. W. B. Maxwell was on lookout for Navajos. With the declaration of martial law and the order to concentrate settlers in large towns, the outlying ranchers and people from the smaller villages began to move into Toquerville, Virgin and Rockville.
  • 26. 172 In June, General Snow decided to abandon Long Valley. Mrs. John Dennett, who made the trek as a girl, recalled the line of wagons leaving Long Valley with armed guards in front and rear. While crossing the sand hills between there and Kanab, a small boy was run over and killed. A halt was made while the child was buried in the sand, but the exigencies of the situation forbade longer delay and the weeping mother was hastily torn from the fresh grave. The settlers’ train passed Kanab to the left and pushed on toward Pipe Springs. Near the mouth of Cottonwood Canyon (June 27) they ran into an ambush of Indians who, for some unexplained reason, failed to attack. J. M. Higbee reports [72] that they called to the Indians to come in and talk or be shot. They came in and talked. According to Mrs. Dennett, there were seven or nine Indians taken into Pipe Springs for a council of war. The wagons were driven into a large circle, as was customary in times of danger, the Indians inside the circle in the center of the group of whites. Higbee says the Indians were told that if any more were found along the route of the caravan they would be shot. Mrs. Dennett adds that some of the Indians had guns and clothes belonging to the Berry boys, which greatly enraged a brother of the dead men, who pleaded to be allowed to revenge his kin. After this, no more Indians were seen on the trip. In the late summer of 1866, Captain James Andrus [73] was ordered to investigate Indian routes crossing the Colorado River in the rough country between the Kaibab and the mouth of the Green River. A group of men was mustered into service from the Virgin River settlements at Gould’s Ranch, twenty-six miles east of St. George, August 16, and moved on to Pipe Springs two days later, where final preparations were made. On the 21st, forty-six mounted men, each equipped with a rifle and two pistols, and with a pack horse bearing forty days’ rations for each pair, started northeast toward the rough country. They went via abandoned Kanab and Scutumpah to the Paria River six miles above Paria settlement where
  • 27. they met another contingent of their party. Two days later, Joseph Fish with eighteen men arrived from Iron county. They located the Ute trail which passed down the Paria to the Colorado. Elijah Averett, sent back with some of the surplus animals, was killed by Indians in the hills west of Paria. On August 29, the main party went northeast through the hills south of the Aquarius Plateau into a valley where they found wild potatoes growing (hence named Potato Valley, now Escalante Valley). They climbed the Plateau and looked off into the wild country stretching to the mouth of the Green River. Convinced that there was no use in going farther, they retraced their march on September 2, traveled to the northwest corner of the plateau, descended to the Sevier River Valley and reached Circleville. They had been pathbreakers from Paria to this point. From here they returned via Parowan and Cedar City. The settlements were now prepared for attack. An Indian raid was made on John D. Lee’s ranch near Beaver on October 23, 1866, and in November, General Snow learned that the Navajos were concentrating east of the Colorado for new raids on Kane County. Soon a friendly Paiute reported that the Navajos were nearing Pipe Springs. The crops planted in Long Valley had been left in the care of friendly Paiutes when the settlers left. In the fall, the Berry boys and others went back to harvest the best crop that had yet been grown there. It took several trips to haul the produce to the Dixie settlements. During their last trip, Snow received a report of an attack of sixteen Navajos on three white men at Maxwell’s Ranch, in which Enoch Dodge was wounded. Snow sent men to Long Valley and instructed A. P. Winsor to throw an intercepting force between the settlements and the fords of the Colorado, to recover lost stock and find out whether the raiders were Navajos or Paiutes. He was promised that other men would be held in readiness if needed. While this force was on the road, the Long Valley party started home with a wagon train. On October 31, when the teams were spread out
  • 28. 173 doubling up Elephant hall, about nine miles south of Mt. Carmel, Indians attacked near the summit and shot Hyrum Stevens. The pioneers abandoned the train and left everything in the hands of the Indians. Stevens was taken with the others on horseback (with a man behind to hold him in place) around the head of Zion Canyon on a three-day trip over the Old Indian Trail and down over Kolob to Virgin. He survived the ordeal and returned to his home at Rockville, where he lived to a ripe old age. When a rescue party under Captain Sixtus E. Johnson arrived a week later, November 5, they found the wagons unattended, tongues broken and contents scattered. The Indians had taken five yoke of oxen, eleven horses and everything they could carry, including harness, flour and wheat. Four Paiutes, however, had pursued the Navajos and recaptured the cattle and harness. Finding the teamsters gone, Johnson gathered up the livestock that had scattered back along the way to Long Valley. Then a second rescue party under Major Russell from Rockville arrived with the Paiutes who had retrieved the harness and cattle. They took the caravan into Virgin, arriving November 11. On November 26, Major John Steele reported signal fires on the mountain south of Virgin City and General Snow issued an order to establish posts at the mouth of (Black) Rock Canyon sixteen miles southeast of St. George and near Gould’s Ranch, eight miles south of Virgin City. [74] The men at these posts were to serve as guards as well as herders of livestock and were to build stone quarters; the “house to be covered with stone flagging or earth in a manner that it cannot be fired from the outside, with but one door and that heavy and strongly barred, so that one or two men, well armed, may defend themselves against any number of Indians.” Despite these precautions, the Navajos scattered in small bands, easily passed through the military posts, and hid in the mountains north of St. George. On the evening of December 28, word reached
  • 29. 174 Harrisburg from local Paiutes that some Navajos had killed and dried three beeves between Grapevine Springs and Toquerville. Captain J. D. L. Pearce, with fifteen men from Washington, at once took up their trail along Harrisburg Creek toward Pine Valley Mountain but failed to overtake them. In the meantime, on December 28, near Pine Valley, Cyrus Hancock saw three Indians skulking on the range and called to them. The Indians proved hostile and tried to capture him. One seized his horse’s bit and another tried to shoot him with an arrow. He slid off his horse and ran toward Pine Valley, the Indians in pursuit. One of them shot him in the arm with an arrow. He stumbled and fell as they yelled in triumph, but he regained his footing and outran them into the valley. These Indians were thought to have been hiding around the town for two or three days, quietly gathering stock. As soon as discovered, they left with about thirty horses and passed down the Black Ridge between St. George and Middleton on the night of December 28, gathering more horses at both places and hastening southwest via Fort Pearce Wash. Col. D. D. McArthur immediately ordered out all available cavalry in pursuit of the thieves, who had an entire day’s start. An expedition of thirty men headed by Lt. Copelan followed the Indian trail from the Washington Fields past Fort Pearce, through Black Rock Canyon and out toward Pipe Springs where it met another detachment returning from an Indian encounter. Captains J. D. L. Pearce and James Andrus were at Harrisburg on the evening of December 29, when an express carrying instructions to Colonel Winsor at Rockville arrived. Upon reading the instructions they started for Rockville and arrived at dawn. Thirty men gathered and pushed on to Maxwell’s ranch where they arrived that evening. After resting an hour, they hastened on to Cedar Ridge and five miles southeast of Pipe Springs. Sixtus E. Johnson spotted the smoke of Indian fires curling up in the distance, about half a mile from the place where Whitmore and McIntyre had been killed. The men
  • 30. 175 slipped into a wash and kept out of sight until within gunshot of the Indians, when they made a dash to get between them and their horses. Firing opened and the Indians took to the rocks. The skirmish lasted nearly an hour and covered a rough area half a mile wide and three miles long. The thirteen Navajos in the band refused to yield even when cornered, and several died fighting. During the fray an arrow aimed at Captain Andrus struck his horse in the forehead, saving the rider. One mortally wounded Indian continued to shoot until he fainted. Another, wounded in both legs, fired until his arrows were spent and then kept twanging his bow as if shooting as long as the fray lasted. When all was quiet the whites gathered together and found that none was injured. Two Indians who had escaped came out on a hill some distance away where they felt safe and slapped their seats in derision and yelled “Squaw! Squaw!” in defiance. A man named Warren, from Pine Valley, who had an extra long range breechloading gun took a chance shot and brought one of them down. The other fled. Of the thirteen Indians, four were killed, seven wounded and two escaped, only one on horseback. Three horses were lost, but the balance and the thirteen cattle were recovered and brought back. Copelan’s party returned on January 1, 1867 but Pearce and Andrus tarried two or three days longer. General Snow was in Salt Lake City at the time and his responsibility fell largely upon Captain J. D. L. Pearce and Adjutant Henry Eyring, his assistant. The concentration in the larger towns and the military control of the movements of people in the region tended to reduce the danger to the settlers. Tension with the local Paiute Indians was gradually eased, although the Navajo raids continued for several years. Jacob Hamblin, Utah’s “Leatherstocking,” played an important role in quieting the Paiutes. In the fall of 1867, he was instructed to keep in touch with the Indians and do his best to pacify them. He went to Kanab, where he helped them plant corn and vegetables and had peace parleys with them, urging the Paiutes to
  • 31. cooperate in preventing Navajo raids by watching the fords of the Colorado and the trails leading to the settlements. In November, 1868, a band of about thirty Navajos crossed the river on foot on a marauding expedition. They divided into squads of two or three and worked at night in different quarters so rapidly as to baffle the pickets. They got away with some stock, although twenty- seven horses were recovered from them on the 25th at Black Canyon, by Andrus and his command. Notice of their presence came on November 22 from Henry Jennings to Erastus Snow at St. George. The next day local Indians reported tracks around St. George, and General Snow ordered the livestock along the Virgin gathered together and herded under armed guard. He placed pickets along the river for fifteen miles and sent Col. J. D. L. Pearce with a company of cavalry to guard the rough country passes from Black Rock Canyon (25 miles southeast of St. George) to Pipe Springs. Two days later, word came from Washington that the Navajos had made off with a band of horses via Black Rock Canyon. On the night of the 26th, a party of these Indians with about twenty horses eluded the guards not far from Pipe Springs and made their way eastward. A detachment under Captain Willis Copelan started in pursuit. He chased the Navajos and was about to overtake them, but before he attacked, some friendly Paiutes encountered the Navajos, gave battle and killed two. They recovered the horses and willingly turned them over to Copelan on his arrival. The Paiutes were rewarded with suitable presents. By December 1, 1868 Pearce concluded that the Navajos had decamped, and started home, moving from Pipe Springs to Cedar Ridge. On that same day, however, Erastus Snow received word from J. W. Young on the Muddy River in Nevada, sixty miles below St. George, that the Navajos had run off with eighteen horses and mules. Snow sent word to Pearce to be on his guard. A posse of whites and Paiutes set out in pursuit from Mesquite. The Indians overtook the Navajos and recovered eleven of the horses.
  • 32. 176 The messenger carrying this news reached Col. Pearce at Cedar Ridge at 4 a.m., December 2, and at daybreak scouts were sent out. Captain Freeman found their trail and started after them with several men, being joined by Captain Copelan. They sighted the Navajos’ dust, but could not overtake them and the chase had to be abandoned. A raid in February 1869 caused such concern that another expedition (February 25 to March 12) of thirty-six men, under the leadership of Captain Willis Copelan was sent out to deal with it. [75] As usual, the Navajos struck swiftly and fled before the expedition arrived. At Pipe Springs, Copelan watched the passes, hunted the surrounding region for the raiders and found they had gone east. On March 1, with twenty men he started in pursuit of the raiders. About eight miles out he struck a trail where the Indians had been driving about fifty head of cattle. During the next five days he followed the trail around the north end of Buckskin Mountain (Kaibab) across Paria and Warm Springs Creek to the old Ute ford on the Colorado River. Finding the quarry had escaped, he returned home, arriving at St. George March 12. During the fall, fresh raids by the Navajos created yet another scare. A band raided settlements north of St. George and drove off stock. This time Colonel James Andrus was detailed to lead a foray against the marauders. He started up the Virgin River gathering fourteen recruits. Then he went to Pipe Springs where he received word that another band of Navajos had raided near Pinto. He hastened toward Pinto to intercept them, passing via Kanab and Scutumpah. Near Paria, he found a trail where some Navajos had escaped with an estimated eighty head of livestock. Here Andrus learned that the Paiutes had attacked and wounded a Navajo in a running fight, and that other raiders were on the way back from Pinto. Andrus and his men waited until November 10 and finding no signs of the Indians, started home. The detachment had not gone far when they encountered a fresh Navajo trail made by an estimated twelve
  • 33. 177 horses and two men. They caught up with the Indians early the next day just as they were passing into a narrow gorge of the Paria canyon. There were actually eight Indians with twelve horses, traveling leisurely. Under the detachment’s fire, two Navajos fell; the rest disappeared into the narrows. A few minutes later they re- appeared on the canyon cliffs on both sides of Andrus’ force. Bullets from the Indian rifles soon convinced Andrus that discretion was the better part of valor and he retired. The Navajos were adroit raiders. In rounding up stock they would often camouflage themselves with bush foliage, crawling past the unsuspecting guards to stampede the herd. Or they would skin a young steer, leaving hoofs and horns in place and throw the hide over a brace of Indians, who would steal to the corral under cover of darkness, let down the bars, and quietly drive the stock away. These raids were costly. Not only did the settlers live in constant fear, but a heavy toll of livestock, estimated in 1869 at 1200 horses and cattle, was taken. Men had to be continually on the alert and peaceful pursuits were interrupted to furnish posses to chase the Indians. When Major J. W. Powell of the U. S. Geological Survey was exploring the Kanab region in 1870, he expressed grave concern about the losses the Mormon settlers were suffering because of the raids. [76] In October, Jacob Hamblin decided to accompany Major Powell on a peace mission to the Navajos when the latter was leaving to return to Washington, DC. [77] They reached Fort Defiance in eastern Arizona at a time when 6,000 Navajos were gathered there for their annual allotments from the Federal government. All the Navajo chiefs but one were present and met in council to consider Hamblin’s proposal. Powell introduced Hamblin by saying that he represented the Mormons from the other side of the Colorado River who were helping to pay the taxes from which the annual allotments to the Navajos were made. Hamblin, in turn, pointed to the disastrous consequences of the war and the advantages of peace.
  • 34. 178 Through war the Navajos had lost twenty or thirty men; with peace they could herd their livestock in distant places where forage was good without fear of molestation. He proposed, in place of war, a peaceful settlement of difficulties and trade with the Mormons. After several days of consultation, peace was agreed upon. The council appointed one of the chiefs, Hastele, who lived near the Colorado River, as negotiator who ended by saying, “We hope we may be able to eat at one table, be warmed by one fire, smoke one pipe and sleep under one blanket.” Thus was peace promised, though it was soon again to be put in jeopardy. Hamblin reached Kanab with the good news about December 11, 1870. [78] Within a few weeks, a group of eighty Navajos arrived at Kanab on a trading expedition. They came on foot and brought all the Navajo blankets they could carry. They scattered among the settlements and traded their blankets for horses and returned well satisfied with the experiment. Peaceful trading continued until the winter of 1873-74, when a party of four young Navajos was caught in a snowstorm near a ranch in Grass Valley, Sevier County. They made themselves at home at the ranch and even killed a small animal for food. The owner of the ranch, said to be a non-Mormon, learned of their presence and gathered some of his friends to go with him to investigate. At the ranch, they shot and killed three of the Indians and wounded the fourth, who escaped and after painful hardships made his way home. [79] His story inflamed Navajo vengefulness and disquieting reports reached the Mormons of threatened reprisals. Brigham Young asked Hamblin to visit the Navajos again and satisfy them that the Mormons were not involved in the outrage. Bishop Levi Stewart of Kanab, however, tried to dissuade Hamblin and even sent a messenger to induce him to return after he had started, urging that the risk was too great.
  • 35. Firm in his purpose, Hamblin went his way and met the Navajos east of Moencopi, about January 29, 1874. Hastele, the representative appointed by the Navajos, was not there, but other influential Indians considered Hamblin’s statement. The war council was held in a Navajo hogan, to which there was but one entrance opposite Hamblin and his two companions, while two dozen Navajos occupied the space between. Hamblin’s explanation of the killings was at first rejected on the ground that it was he who had invited the Navajos to come into the Mormon country to trade, with the result that three of their good young men now lay on the ground “for the wolves to eat.” The interpreter told Hamblin his companions could go home, but he must die. The moment was tense. His companions refused to leave him. Without arousing suspicion, Hamblin passed several revolvers to his friends, saying as he did so, “These are in my way.” The men behind unobtrusively readied them in case of emergency. Hamblin reminded the Indians of his many friendly acts, of his willingness to come into their midst to settle the matter, and told them it was not right to kill him for the acts of strangers for whom he was not responsible. The wounded Indian was brought in. A stirring appeal for revenge was made by a young warrior, who demanded that Hamblin be the victim. The Indians, however, after the excitement subsided, offered to settle for three hundred and fifty horses and cattle. Hamblin deliberately refused. One of them remarked that he would agree after he had been stretched over the hot coals of the fire. The interpreter asked if he were not afraid. “No,” he said, “my heart has never known fear. What is there to scare me?” “The Navajos,” was their answer, to which he replied that he “was not afraid of his friends.” Mollified, the Indians finally agreed to leave the matter to be settled by Hastele after an investigation. Late that spring, Hastele and his party visited Kanab and were piloted to Sevier Valley where their findings convinced them that the Mormons were innocent. Thus ended the last threat to peaceful
  • 36. 179 relations with the Navajos. Thereafter, both groups traded on good terms largely due to the outstanding bravery and cool judgment of Jacob Hamblin.
  • 37. Expansion in Kane County Re-settlement of Long Valley and Kanab does not seem to have been attempted until 1870, although Kanab and Paria were occupied by missionaries under Jacob Hamblin in 1867 as frontier outposts. At Paria a strong guard house and corral was built and some land was cultivated, beginnings out of which the settlement grew with the accession of several families in 1872 and 1873. Kanab was similarly restored. The necessity of a fort there was impressed upon the whites by the continued Navajo raids. Five stone masons were sent from St. George in 1869 to construct the fort. They reached Kanab on August 28 and worked until early in September, when John R. Young told them they had finished their mission and could go home. This building expedition brought new settlers to Kanab, for John Mangum (or Mangram), his brother, James, James Wilkins, and George Ross, moved there soon after. Nate Adams, who visited Kanab in September, 1870, and who moved there March 14, 1871, says the first three were in hiding and that John D. Lee, also in hiding, took up Scutumpah Ranch and explored Lee’s Ferry in 1869. Several missionaries were sent to aid Hamblin about the same time. They were fencing and cultivating land when Brigham Young made his first visit to the Kanab country about the 1st of April, 1870. George Albert Smith wrote of this visit: At Kanab we met Brothers Jacob Hamblin and Jehiel MacConnel [McConnell], and several other missionaries, who were engaged in teaching the Indians how to cultivate the soil and to obtain a living by peaceful pursuits. We were much pleased with the country.... As soon as measures shall be taken to prevent the annual raids of the
  • 38. 180 Navajos, this land of Canaan will be re-occupied by the Saints and become a valuable acquisition to our southern settlements. [80] A pioneer Salt Lake photographer, C. R. Savage, took many pictures along the way, including one of Brigham Young and his party on the Colorado at the mouth of the Virgin, and several of Zion Canyon. Upon his return to Salt Lake City on April 16, 1870, Brigham Young sent a group of fifty-two people led by Levi Stewart, to re-settle the Kanab country. They went down through the Dixie settlements and reached Pipe Springs on June 1. They remained there and at Moccasin Springs several days while exploring the region. [81] On the 14th they moved over to Kanab Creek and joined Jacob Hamblin at the old fort, now too small to house so many. Brigham Young manifested much interest in the success of the colony. He promised Stewart that he would visit him in the fall and asked him to find a more direct route to Kanab from the north that would obviate the long roundabout approach through the Dixie settlements and the Arizona strip. Stewart sent out two expeditions, the second of which found a road from the head of the Sevier River through Upper Kanab and Johnson Wash. Brigham Young started for Kanab on August 26, 1870. Accompanying him from Parowan was the intrepid explorer and topographer of the U. S. Geological Survey, Major J. W. Powell, who had already made one trip through the Grand Canyon and was returning to make plans for further geological studies and his second trip through the canyon. [82] In attempting to follow Stewart’s directions, the party lost its way and wandered into the Paria River valley and thus went many miles out of its direct route. According to Nate Adams, [83] “old Humpy Indian” guided the company safely into Kanab on the evening of September 9, 1870. While there, a townsite and fields were surveyed
  • 39. east of the fort beyond the path of the canyon winds. Brigham Young returned to Salt Lake City via St. George and the Dixie settlements. Three months later, on December 14, 1870, six lives were lost in a fire at the fort. These included Bishop Stewart’s wife, Margery, and three of his sons. [84] Brigham Young made a special trip to Kanab from St. George, where he was wintering, to comfort the bereaved families. Soon after, the settlers began to build their homes on the townsite. Within a few years, the fort was deserted but it was maintained for some time for use in case of emergency. Dellenbaugh, a member of Major J. W. Powell’s party, thus describes his visit to Kanab in the early 70’s:
  • 40. F. S. Dellenbaugh in Zion Canyon (1930) with the flag of the Emma Dean boat that made “a canyon voyage” about 60 years
  • 41. Photo U. S. Nat. Park Service. Courtesy Union Pacific Railroad. 181 earlier. Viewing Bryce Canyon from the rim. ... Nigger, [a white mule] went along very well and I was in Kanab by three o’clock. The village which had been started only a year or two, was laid out in the characteristic Mormon style, with wide streets and regular lots fenced by wattling willows between stakes. Irrigating ditches ran down each side of every street and from them the water, derived from a creek that came down a canyon back of the town, could be led into any of the lots, each of which was about one quarter of an acre; that is, there were four lots to a block. Fruit trees and vines had been planted and were already beginning to promise near results, while corn,
  • 42. 182 potatoes, etc., gave fine crops. The original place of settlement was a square formed by one-story log houses on three sides and a stockade on the fourth. This was called the fort and was a place of refuge though the danger from Navajo attack seemed to be over and that from any assault by the Paiutes certainly was past. One corner of the fort was made by the walls of the schoolhouse, which was at the same time meeting-house and ball-room. Altogether there were about 100 families in the village. The houses that had been built outside the fort were quite substantially constructed, some of adobe or sun-dried brick. The entire settlement had a thrifty air, as is the case with the Mormons. Not a grog-shop, or gambling saloon, or dance-hall was to be seen; quite in contrast with the usual disgraceful accompaniments of the ordinary frontier towns. A perfectly orderly government existed, headed by a bishop appointed by the church authorities in Salt Lake City, the then incumbent of this office being an excellent man, Bishop Stewart. [85] After the Navajo peace settlement many of the places abandoned in 1866 were reoccupied and within a few years further expansion filled most of the remaining areas suitable for settlements or ranches. In Long Valley, Berryville (now Glendale), and Winsor (Mt. Carmel) were revived in 1871. Johnson was settled in the spring of the same year by five brothers, Joel, Benjamin, Joseph, George and William Johnson, on the site of Scutumpah, formerly John D. Lee’s ranch. In 1872, Graham, on the headwaters of Kanab Creek (upper Kanab), was reoccupied and the settlers engaged in dairying and lumbering. The upper reaches of the Paria, however, attracted settlers from the north. Panguitch was re-founded in March, 1871 under George W. Sevy and counted seventy-five families the next year. Joel H. Johnson and George D. Wilson established a sawmill in 1871 near the present location of Hillsdale, and were soon joined by twenty families, including those of Nephi and Seth Johnson. Other cattlemen located farther up the Sevier, where Meltiar Hatch founded the village bearing his name. Nephi Johnson, discoverer of Zion Canyon, was made bishop of Hillsdale in 1874.
  • 43. Attention was then focused on the upper Paria. The first settlers, David O. Littlefield and Orley D. Bliss, located near the present site of Cannonville on Christmas Eve, 1874. Early the next day eight other families arrived, who built log houses at a place called Clifton and began farming along the Paria and on Henrieville Creek. Ebenezer Bryce, from Pine Valley, selected a place farther upstream, a mile or two east of the present site of Tropic near the mouth of Bryce Canyon. Bryce used the famous canyon for a cattle range, and thus immortalized his name. Clifton was not well located and in 1877 some settlers moved to a new townsite called Cannonville, in honor of George Q. Cannon, high Mormon official who had taken a special interest in their affairs. Other settlers moved over to Henrieville Creek to be near their farms, and thus the town of Henrieville (named for James Henrie, president of Panguitch Stake) was born. In 1879 Daniel Goulding settled near Bryce’s ranch. Seeking water for irrigation, he and Bryce devised a scheme to divert water from Pine Creek in the Great Basin by means of a canal over the divide. This they finished, but upkeep was expensive, their crops were poor, and Goulding lost about five hundred fruit trees from drouth. Bryce became discouraged and left for Arizona in 1880 and Goulding moved to Henrieville in 1883. Bryce, unimpressed by the beauty of the canyon, considered it “awful hard to find a cow that was lost” in the intricate maze of its pinnacles. Seth Johnson and several others located in 1886 on Yellow Creek (Kane County) about three miles southwest of Cannonville and named the settlement Georgetown, in honor of the same man for whom Cannonville was named. In 1890 the two Ahlstrom brothers built homes on the present site of Tropic and with several others began a second and more ambitious attempt to divert water from the East Fork of Sevier River over the divide into Paria Creek. This time the project succeeded. Tropic townsite was surveyed in 1891 and settlers began to flock there and
  • 44. 183 prepare homes and lands in anticipation of the coming of the water. A fitting celebration was staged on May 23, 1892, when the water was turned into the canal. By this time, most of the suitable valleys and canyons had been occupied. Erosion, however, caused trouble at Kanab. From 1883 to 1890, floods presumably resulting from overgrazing tore out dams and ditches and gutted the canyons and valleys with deep washes. Water arose in the bottom of the washes and that in Kanab Wash (below Kanab) was diverted about 1886 onto a new townsite just beyond the state line in Arizona, called Fredonia, which later served as a refuge for a number of polygamous wives during the Federal offensive against the practice. [86] While southern Utah was thus growing, a new movement was developed. In 1879, the Mormon Church leaders called for eighty men from the Southern Mission to establish an outpost for the purpose of “cultivating and maintaining friendly relations with Indians whose homes were near the point where the state of Colorado and the Territories of Utah, New Mexico and Arizona come together.” Twenty-five men, including Kumen Jones, went out to investigate routes and locations. They traveled via Lee’s Ferry, Tuba City and Monument Valley to the San Juan River as far as Four-Corners, spent about three months exploring the region, and then returned home via a northern route, past the sites of Monticello and Moab. [87] While they were away, another party set out from Escalante seeking a short-cut to the San Juan country. A route much more direct than that mapped by the first party was reported, apparently on imperfect observation. It was, however, accepted, and by October the party was on its way. It passed through Escalante and reached Forty-mile Spring where it was held up by excessively rough country, while snows in the mountains blocked retreat. Three scouts were sent ahead to investigate some of the wildest and most rugged scenic areas of America. The three returned in
  • 45. 184 disappointment; one held the route feasible, another positively rejected it, while the third thought it might be possible to get through with special help. Envoys were sent to Salt Lake City to appeal for assistance, which was given in the form of a legislative appropriation for blasting a way through. It took fifty days to get eighty-two wagons through Hole-in-the-Rock and down to the Colorado River and ninety days to reach Bluff on the San Juan River where the first settlement was made. Three babies were born on the way and the hardships endured form a Western epic. [88] The story of Orderville has been left for the last. The United Order [89] was organized at Mt. Carmel, March 20, 1874, by John R. Young, at which time one hundred and nine members were listed. One summer of the United Order was enough for most members. Bishop Bryant Jolley, with his numerous family and relatives, formed the core of the dissenters. To avoid contention, those who wished to continue with the Order sold their holdings and moved in a body two miles above Mt. Carmel where title to all land was vested in the group and where they set up the town Orderville, under the leadership of Howard O. Spencer. The new town was surveyed February 20, 1875. The first building was a hotel where all ate together in the large dining hall, from July, 1875 to May, 1880. As time passed, living quarters were provided for each family and work was divided into more specialized fields. During the hey-day of the Order, around 1880, it numbered nearly six hundred adherents and there were some twenty-eight specialized departments of work which included most of the various activities that go to make up a simple community. The Order made great progress and acquired property rapidly. Farming lands were expanded to include areas scattered through Long Valley and at Kanab.
  • 46. 185 The growing power of the Order created jealousies, but disintegration came from internal dissension. The idea of giving everyone an equal reward regardless of ability or accomplishment tended in many cases to lessen effort and brought charges of laziness and carelessness. Gradually, more and more individual property was assigned to each home; farmers were given a share of their own produce and livestock and sawmills and freighting were operated under lease or contract. Matters were hastening toward dissolution when, in 1885, polygamy troubles began. Fear that the Federal government might confiscate the goods of the Order forced the final dissolution of most of the property, and farming lands, livestock, ranches, tannery and sawmill, were all sold to members. The woolen mill alone was kept and intermittently operated until 1900. In that year the United Order of Orderville was officially dissolved, twenty-five years after its incorporation.
  • 47. Zion Canyon “In an instant, there flashed before us a scene never to be forgotten. In coming time it will, I believe, take rank with a very small number of spectacles each of which will, in its own way, be regarded as the most exquisite of its kind which the world discloses. The scene before us was the Temples and Towers of the Virgin.” Thus prophetically wrote Captain Clarence E. Dutton of the U. S. Geological Survey in a report published in the year 1880. Dutton was following up the geological work begun by Major J. W. Powell ten or twelve years earlier when the latter started out to explore the Colorado River and made his two memorable trips in boats down the river through the Grand Canyon. The geological problems encountered were so extraordinary that Dutton was detailed to further investigation and encountered problems that have engaged the attention of scientists to this day, particularly the eminent geologist, Dr. Herbert E. Gregory. Following the line of Vermillion Cliffs from Kanab westward, Dutton came to the pass between Eagle Crags and Smithsonian Butte when suddenly, startlingly, there lay before him to the northward the valley of the Virgin River in all its grandeur. Few have seen Zion as Clarence Dutton saw it. From a high pass, in late afternoon, with the sun on his left, he looked into that vast panorama of the Vermillion Cliffs of Zion and Parunuweap and those flanking the Great West Canyon as well—a twenty-mile stretch in one sweeping view. The setting sun cast shadows that made the turrets and towers stand out in bold relief, while the light reflected from one wall upon another intensified the tints and shades of the reds until they stood out in striking contrast with the vivid green of the
  • 48. 186 vegetation and the higher cliffs. No wonder the cold scientist broke down and described in emotional terms this superb panorama. Forerunners of Dutton had visited Zion, but none had penned such eloquent praise. Major J. W. Powell and two companions, Stephen V. Jones, one of his topographers, and Joseph W. Young, a Mormon, left Long Valley on September 10, 1872 [90] and started down through the Parunuweap on foot. They came out next day before noon and spent another day visiting Zion Canyon. Of this trip, Powell says of the Parunuweap: At noon, we are in a canyon 2500 feet deep and we come to a fall where the walls are broken down, and the huge rocks beset the channel, on which we obtain a foothold to reach a level two hundred feet below. Here the canyon is again wider and we find a floodplain on which we can walk. Next day of the Mukuntuweap Canyon, he writes: Entering this, we have to wade up the stream; often the water fills the entire channel, and although we travel many miles, we find no floodplain, talus, or broken piles of rock at the foot of the cliff. The walls have smooth, plain faces, and are everywhere very regular and vertical for a thousand feet or more, where they seem to break in shelving slopes to higher altitudes; and everywhere as we go along, we find springs bursting out at the foot of the walls. [91] Jack Hillers, a photographer from Powell’s party, spent some time during the summer of 1873 taking pictures in Zion Canyon. These are on file in the U. S. Geological Survey Office and have been often used in publications. For a long time, however, this material and Zion Canyon were largely forgotten. The local course of development continued placidly for many years. Only occasionally a hardy traveler, hearing of the beauties of the region, had courage enough to brave the rocky, dusty roads to enjoy the scenic splendors. One was Frederick S. Dellenbaugh, who had accompanied Powell on his
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