![Translated From Etudes Religieuses, Etc.,
Par Des Peres De La Compagnie De Jesus.
The Flight Of Spiders.
A Paper Read Before The French Academy Of
Science, March, 1867.
About fifteen years ago, I was sitting in an arbor of my garden,
reading, when a little spider fell on my book, whence I could not
tell, and commenced to run over the very line I was reading. I blew
hard to chase him away, but he would not go. He lifted himself
strangely up, and I cannot explain how, but he lodged on a sprig of
verdure just above my head. "Well," said I, "for a little animal like
that, this is a wonderful feat! How has he accomplished it?" To
satisfy myself, I took him up again, balanced him on my book, and,
after assuring myself that he had no invisible thread to aid him, I
blew again, and again the little fellow did the very same thing. With
redoubled curiosity, I tried him once more, and, to see better, I sat
down in the bright sunlight. Again I balanced him on the book,
looked at him as closely as possible, and, when I felt assured no
precaution could have escaped me, I blew once more. ... Resuming
the same inclined position, the spider as quick as lightning darted
the finest possible thread out of him, raised himself in the air, and
disappeared.
I confess I was stupefied. Never had I imagined these little animals
could fly without wings; so I consulted several works on zoology,
but I was astonished to find there was no mention made of the
flight of spiders, nor of the ejaculatory movement of which I had
witnessed so curious an example. [Footnote 96]](https://image.slidesharecdn.com/98820-250308211539-f38eed34/85/DNA-Nanotechnology-Methods-and-Protocols-1st-Edition-Chenxiang-Lin-37-320.jpg)
![[Footnote 96: In M. Eugène Simon's Natural History of
Spiders, the most recent work of the kind, he says,
speaking of the manner in which l'épéire diadème
constructs its web: "Several authors suppose that the
spider darts its thread like an arrow, others imagine it
throws it upward in the air while flying as a fly would;
but neither of these explanations rests on observation,
and they are, after all, simple hypotheses." Then,
describing his own observation as to how a spider acts
to make fast its great threads, he says, "It seems to
take a horizontal position, and moves contrary to the
wind." M. Simon's work gives us nothing else to lead us
to suppose he has observed the wonders spoken of.—Tr.]
So there was a new question presented to me, and my vocation to
study the habits of these little animals—which hitherto had given
me no concern—decided for me. I immediately lost all repugnance,
all distaste, and threw away all the unjust precautions of which the
spider is too often the object, and of which I was as culpable as
any one else. And from that time I welcomed its appearance; was
most happy to meet with it, looked for it, indeed, and studied its
habits almost with furor. And I can say that, thanks to this hearty
preoccupation, which never left me, I found every opportunity to
follow my inclination, and knew where to find spiders in all sorts of
unheard-of places.
Such are the singular effects of curiosity once excited, and still
another proof that, in order to study nature well, we need only a
mysterious glimpse of the unknown to redouble all our energies to
explain it thoroughly.
And as in this study, trifling as it may appear, I seem to have met
with facts not known hitherto, but which deserve to be understood,
I here resume the principal ones: those that treat of the flying of
spiders; of the habitation of some species in the air; and of the
gossamer or air threads—a singular phenomenon, for a long time](https://image.slidesharecdn.com/98820-250308211539-f38eed34/85/DNA-Nanotechnology-Methods-and-Protocols-1st-Edition-Chenxiang-Lin-38-320.jpg)
![concerning them any responsibility. But I was firmly convinced, and
therefore hoped to be believed, especially by this generation of
fearless naturalists, who are astonished at nothing in nature, and
who, having often been surprised in the relation of almost
incredible marvels, must certainly make allowances for a few more
in another quarter.
Let us look at, for instance, the wonderful things related of the
argyronete, or aquatic spider. [Footnote 97]
[Footnote 97: The argyronete is a spider that lives in
the water where she constructs a charming little edifice
that appears surrounded with a silky mortar. The down
that covers her contains a certain quantity of air for
respiration. This gives her in swimming the appearance
of a ball of quicksilver, from which we have her name.]
I could not tell anything more unlikely, so I will only exact for the
atmosphere a companion to what the Père de Lignac discovered in
the last century for the water. Yes, I pretend there are spiders that
live in the air as well as those living in water, and that every year,
from the earliest days of spring, there is, unknown to us, a general
migration of spiders toward the atmosphere, where they pass their
best season, form their nets, chase their prey, and only return to
earth in the first fogs of autumn to find their quarters for the
winter. I add, also, that this ascent and descent give rise to the
curious phenomenon, still so badly explained, of the gossamer. And
as it was to the study of this phenomenon that I owe my
knowledge of the rest, may I be permitted here, by way of
demonstration, to relate briefly the path I have followed and the
proofs which have led to the conviction I express?
Attracted, as I was, by all that concerns spiders, I could not remain
indifferent to a fact so important and interesting as the periodical
apparition of those threads which in spring and autumn we see
flying about in long white skeins, clinging to trees, to hedges, and](https://image.slidesharecdn.com/98820-250308211539-f38eed34/85/DNA-Nanotechnology-Methods-and-Protocols-1st-Edition-Chenxiang-Lin-43-320.jpg)
![walked in the fields, the meadows, the gardens, on the borders of
the woods, among heaps of dried leaves, scattered all through the
forest everywhere, could be seen myriads of these little brown
spiders, jumping up and flying before me in every direction, and
exactly such as I had already recognized as such excellent
swimmers. After having passed the winter in the earth, in the holes
of worms that they completed with a little silk, they reappeared
after the cold in great numbers, to disappear again entirely in the
first bright days of spring, and as if by enchantment. If one is seen
again during the summer, we may be sure it is some female
retarded by laying her eggs, and dragging laboriously her cocoon
after her. Now, what has become of the others?
For several months I could not satisfy myself on this point, when,
on the 21 St of October, 1856, in the enclosure of the little
seminary of Iseure, near Moulins, I came to a positive decision, I
was observing the fall of a large quantity of gossamers, which were
falling on that day in large white flakes, when I perceived close to
me in the air one of those little black spiders descending gradually,
and as if she were jumping. She held by an invisible thread to a
large flake, which came down slowly about seven or eight yards
above her; but, keeping outside of it, she hung by the end of the
long thread, like an aeronaut underneath his balloon. My attention
once attracted, I noticed so great a number that I was astonished I
had not taken care sooner; for there was scarcely a flake
underneath which there were not one or two, and this sometimes
even before the flake itself was visible. [Footnote 98]
[Footnote 98: There is an observation which confirms my
own. We read in Darwin's Journal, page 159: "Mr.
Darwin saw a large number of gossamers on the ship
Beagle, when she was about 60 miles from the mouth of
the Rio de la Plata. It was the first of November, and
these gossamers were carried by a very light breeze,
and on each were found an immense number of little
spiders, similar in appearance, about the twelfth of an](https://image.slidesharecdn.com/98820-250308211539-f38eed34/85/DNA-Nanotechnology-Methods-and-Protocols-1st-Edition-Chenxiang-Lin-45-320.jpg)
![inch in length, and in color a deep brown. The smallest
were a deeper shade than the others. None were found
on the white tufts, but all on threads." Journal of
Researches into the Natural History and Geology
of the Countries visited during the Voyage of his
Majesty's Ship, the Beagle, 1845.]
Each one was separated by a slender thread, and followed the
motion of its balloon. If they met a tree or a bush, they landed
upon it; if not, coming close to the earth, they ran along and were
lost in the verdure. If I approached them too quickly or made a
noise, they remounted rapidly by their threads and went to
disembark somewhere else.
I also examined some of the flakes. They were all shining white
mats, appearing as if they had been washed. Several contained
wings and feet of flies, fragments of the case of little coleoptera,
and other remnants of their aerial festivities.
This encounter was for me a revelation. I knew where the spiders,
whom I had seen disappear so brusquely, took refuge, and,
however rash my judgment may appear, I felt assured I had solved
an interesting problem.
But to establish seriously and give to science an opinion so new
and original as that the atmosphere may be peopled with spiders, I
soon felt that more proof was necessary in order to sit down calmly
under my personal conviction. So I concluded I should not be doing
too much if I added to the verification of their descent that of their
ascension, and could surprise them in this new migration. I waited,
therefore, impatiently for the spring.
But that spring, and for five or six that followed it, great was my
disappointment; for, though I perceived several isolated ascensions,
yet nothing in the proportion I had imagined or that could justify
my hypothesis. I began then to doubt seriously my success, when](https://image.slidesharecdn.com/98820-250308211539-f38eed34/85/DNA-Nanotechnology-Methods-and-Protocols-1st-Edition-Chenxiang-Lin-46-320.jpg)
DNA Nanotechnology Methods and Protocols 1st Edition Chenxiang Lin
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- 5. DNA Nanotechnology Methods and Protocols 1st Edition Chenxiang Lin Digital Instant Download Author(s): Chenxiang Lin, Yonggang Ke, Rahul Chhabra, Jaswinder Sharma, Yan Liu (auth.), Giampaolo Zuccheri, Bruno Samorì (eds.) ISBN(s): 9781617791420, 1617791423 Edition: 1 File Details: PDF, 10.00 MB Year: 2011 Language: english
- 7. Me t h o d s i n Mo l e c u l a r Bi o l o g y ™ Series Editor John M. Walker School of Life Sciences University of Hertfordshire Hatfield, Hertfordshire, AL10 9AB, UK For further volumes: http://www.springer.com/series/7651
- 9. DNA Nanotechnology Methods and Protocols Edited by Giampaolo Zuccheri and Bruno Samorì Department of Biochemistry, University of Bologna, Bologna, Italy
- 10. Editors Giampaolo Zuccheri, Ph.D. Department of Biochemistry University of Bologna Bologna, Italy giampaolo.zuccheri@unibo.it Bruno Samorì Department of Biochemistry University of Bologna Bologna, Italy bruno.samori@unibo.it ISSN 1064-3745 e-ISSN 1940-6029 ISBN 978-1-61779-141-3 e-ISBN 978-1-61779-142-0 DOI 10.1007/978-1-61779-142-0 Springer New York Heidelberg London Dordrecht Library of Congress Control Number: 2011929163 © Springer Science+Business Media, LLC 2011 All rights reserved. This work may not be translated or copied in whole or in part without the written permission of the publisher (Humana Press, c/o Springer Science+Business Media, LLC, 233 Spring Street, New York, NY 10013, USA), except for brief excerpts in connection with reviews or scholarly analysis. Use in connection with any form of information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed is forbidden. The use in this publication of trade names, trademarks, service marks, and similar terms, even if they are not identified as such, is not to be taken as an expression of opinion as to whether or not they are subject to proprietary rights. Printed on acid-free paper Humana Press is part of Springer Science+Business Media (www.springer.com)
- 11. v Preface Giorgio Vasari, a painter, architect, and art historian during the Italian Renaissance, is credited with coining the expression “andare a bottega,” (“attending the studio”) refer- ring to the internship that the apprentice would complete in the master’s studio in order to learn what could be uniquely transmitted in person and in that particular environment and that could then lead to making a unique artist of the apprentice. Nowadays, this same concept holds true in science, and despite the many opportuni- ties for communication and “virtual presence”, the real physical permanence in a lab is still the best way for a scientist to learn a technique or a protocol, or a way of thinking. A book of protocols, such as this, humbly proposes itself as the second-best option. Not quite the same as being in person in a lab and witnessing the experts’ execution of a protocol, it still holds many more details and hints than the usually brief methods section found in research papers. This book of protocols for DNA nanotechnology was composed with this concept in mind: prolonging the tradition of Methods in Molecular Biology, it tries to simplify researchers’ lives when they are putting in practice protocols whose results they have learnt in scientific journals. DNA is playing a quite important and dual role in nanotechnology. First, its proper- ties can nowadays be studied with unprecedented detail, thanks to the new instrumental nano(bio)technologies and new insight is being gathered on the biological behavior and function of DNA thanks to new instrumentation, smart experimental design, and proto- cols. Second, the DNA molecule can be decontextualized and “simply” used as a copoly- mer with designed interaction rules. The Watson–Crick pairing code can be harnessed towards implementing the most complicated and elegant molecular self-assembly reported to date. After Ned Seeman’s contribution, elegantly complicated branched structures can be braided and joined towards building nano-objects of practically any desired form. DNA nanotechnology is somewhat like watching professional tennis players: every- thing seems so simple, but then you set foot on the court and realize how difficult it is to hit a nice shot. When you see the structural perfection of a self-assembling DNA nano- object, such as a DNA origami, you marvel at how smart DNA is as a molecule and won- der how many different constructs you could design and realize. Among the others, this book tries to show the procedures to follow in order to repeat some of the methods that lead to such constructs, or to the mastering of the characterization techniques used to study them. Many details and procedures are the fruit of the blending of artistry, science, and patience, which are often unseen in a journal paper, but that could be what makes the difference between a winning shot and hitting the net. Many research groups share their expertise with the readers in this book. For the sake of conciseness, we here mention the group leaders, while it is truly from the daily work of a complete team that the details of a protocol can be worked out. The chapters of this book can be roughly divided into two parts: some deal with the methods of preparing the nanostructures, from the rationale of the operations to the techniques for their handling; some other chapters deal more directly with advanced instrumental techniques that can manipulate and characterize molecules and nanostructures. As part of the first group, Roberto Corradini introduces the reader to the methods and choices for taming helix chirality, Alexander Kotlyar, Wolfgang Fritzsche, Naoki Sugimoto, and James Vesenka
- 12. vi Preface share their different methods in growing, characterizing, and modifying nanowires based on G tetraplexes; Hao Yan and Friedrich Simmel teach all the basics for implementing the self-assembly of branched DNA nanostructures, and then characterizing the assembly. Hanadi Sleiman tells about hybrid metal–DNA nanostructures with controlled geometry. Frank Bier shows the use of rolling circle amplification to make repetitive DNA nanostruc- tures, while, moving closer to technological use of DNA, Arianna Filoramo instructs on how to metalize double-stranded DNA and Andrew Houlton reports on the protocol to grow DNA oligonucleotides on silicon. Also with an eye to the applicative side, Yamuna Krishnan instructs on how to insert and use DNA nanostructures inside living cells. On the instrument side, Ciro Cecconi and Mark Williams introduce the readers to methods for the use of optical tweezers, focusing mainly on the preparation of the ideal molecular construct and on the instrument and its handling, respectively. John van Noort and Sanford Leuba give us protocols on how to obtain sound data from single-molecule FRET and apply it to study the structure of chromatin. Claudio Rivetti teaches the reader how to extract quantitative data from AFM of DNA and its complexes, while Matteo Castronovo instructs on the subtleties of using the AFM as a nanolithography tool on self-assembled monolayers; Jussi Toppari dwelves on the very interesting use of dielectrophoresis as a method to manipulate and confine DNA, while Matteo Palma and Jennifer Cha explain methods for confining on surfaces DNA and those very same types of DNA nanostruc- tures that other chapters tell the reader how to assemble. Aleksei Aksimientev shows the methods for modeling nanopores for implementing DNA translocation, a technique bound to find many applications in the near future. We hope this book will help ignite interest and spur activity in this young research field, expanding our family of enthusiastic followers and practitioners. There are certainly still many chapters to be written on this subject, simply because so much is happening in the labs at this very moment. There will certainly be room for the mainstreaming of pro- tocols on the use of DNA analogues (starting with the marvelous RNA, of course), for the design and preparation of fully 3D architectures, for the development of routes towards functional DNA nanostructures, which will lead to applications. DNA nanostructures can be “re-inserted” in their original biological context, as microorganisms can be convinced to replicate nanostructures or even code them. And eventually, applications will require massive amounts of the nanostructures to be produced and to be manipulated automati- cally, possibly with a precision and output rate similar to that of the assembly of microelec- tronics circuitry nowadays. Our personal wish is that the next chapters will be written by some of our readers. Bologna, Italy Giampaolo Zuccheri Bologna, Italy Bruno Samorì
- 13. vii Contents Preface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . v Contributors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ix 1 Synthesis and Characterization of Self-Assembled DNA Nanostructures . . . . . . . . 1 Chenxiang Lin, Yonggang Ke, Rahul Chhabra, Jaswinder Sharma, Yan Liu, and Hao Yan 2 Protocols for Self-Assembly and Imaging of DNA Nanostructures . . . . . . . . . . . . 13 Thomas L. Sobey and Friedrich C. Simmel 3 Self-Assembly of Metal-DNA Triangles and DNA Nanotubes with Synthetic Junctions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33 Hua Yang, Pik Kwan Lo, Christopher K. McLaughlin, Graham D. Hamblin, Faisal A. Aldaye, and Hanadi F. Sleiman 4 DNA-Templated Pd Conductive Metallic Nanowires . . . . . . . . . . . . . . . . . . . . . . 49 Khoa Nguyen, Stephane Campidelli, and Arianna Filoramo 5 A Method to Map Spatiotemporal pH Changes Inside Living Cells Using a pH-Triggered DNA Nanoswitch . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61 Souvik Modi and Yamuna Krishnan 6 Control of Helical Handedness in DNA and PNA Nanostructures . . . . . . . . . . . . 79 Roberto Corradini, Tullia Tedeschi, Stefano Sforza, Mark M. Green, and Rosangela Marchelli 7 G-Quartet, G-Quadruplex, and G-Wire Regulated by Chemical Stimuli . . . . . . . . 93 Daisuke Miyoshi and Naoki Sugimoto 8 Preparation and Atomic Force Microscopy of Quadruplex DNA . . . . . . . . . . . . . 105 James Vesenka 9 Synthesis of Long DNA-Based Nanowires . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 115 Alexander Kotlyar 10 G-Wire Synthesis and Modification with Gold Nanoparticle . . . . . . . . . . . . . . . . . 141 Christian Leiterer, Andrea Csaki, and Wolfgang Fritzsche 11 Preparation of DNA Nanostructures with Repetitive Binding Motifs by Rolling Circle Amplification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 151 Edda Reiß, Ralph Hölzel, and Frank F. Bier 12 Controlled Confinement of DNA at the Nanoscale: Nanofabrication and Surface Bio-Functionalization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 169 Matteo Palma, Justin J. Abramson, Alon A. Gorodetsky, Colin Nuckolls, Michael P. Sheetz, Shalom J. Wind, and James Hone 13 Templated Assembly of DNA Origami Gold Nanoparticle Arrays on Lithographically Patterned Surfaces . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 187 Albert M. Hung and Jennifer N. Cha 14 DNA-Modified Single Crystal and Nanoporous Silicon . . . . . . . . . . . . . . . . . . . . 199 Andrew Houlton, Bernard A. Connolly, Andrew R. Pike, and Benjamin R. Horrocks
- 14. viii Contents 15 The Atomic Force Microscopy as a Lithographic Tool: Nanografting of DNA Nanostructures for Biosensing Applications . . . . . . . . . . . . . . . . . . . . . . 209 Matteo Castronovo and Denis Scaini 16 Trapping and Immobilization of DNA Molecules Between Nanoelectrodes . . . . . 223 Anton Kuzyk, J. Jussi Toppari, and Päivi Törmä 17 DNA Contour Length Measurements as a Tool for the Structural Analysis of DNA and Nucleoprotein Complexes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 235 Claudio Rivetti 18 DNA Molecular Handles for Single-Molecule Protein-Folding Studies by Optical Tweezers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 255 Ciro Cecconi, Elizabeth A. Shank, Susan Marqusee, and Carlos Bustamante 19 Optimal Practices for Surface-Tethered Single Molecule Total Internal Reflection Fluorescence Resonance Energy Transfer Analysis . . . . . . . . . . . . . . . . 273 Matt V. Fagerburg and Sanford H. Leuba 20 Engineering Mononucleosomes for Single-Pair FRET Experiments . . . . . . . . . . . 291 Wiepke J.A. Koopmans, Ruth Buning, and John van Noort 21 Measuring DNA–Protein Binding Affinity on a Single Molecule Using Optical Tweezers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 305 Micah J. McCauley and Mark C. Williams 22 Modeling Nanopores for Sequencing DNA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 317 Jeffrey R. Comer, David B. Wells, and Aleksei Aksimentiev Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 359
- 15. ix Contributors Justin J. Abramson • Department of Mechanical Engineering, Columbia University, New York, NY, USA Aleksei Aksimentiev • Department of Physics, University of Illinois at Urbana-Champaign, Urbana, IL, USA Faisal A. Aldaye • Department of Systems Biology, Harvard Medical School, Boston, MA, USA Frank F. Bier • Department of Nanobiotechnology & Nanomedicine, Fraunhofer Institute for Biomedical Engineering, Branch Potsdam-Golm, Potsdam, Germany Ruth Buning • Leiden Institute of Physics, Leiden Universiteit, Leiden, The Netherlands Carlos Bustamante • Howard Hughes Medical Institute, Department of Physics, University of California, Berkeley, CA, USA Stephane Campidelli • CEA Saclay, Laboratoire d’Electronique Moléculaire, Gif-sur-Yvette Cedex, France Matteo Castronovo • Department of Biology, MONALISA Laboratory, College of Science and Technology, Temple University, PA, USA Ciro Cecconi • CNR-Istituto Nanoscienze S3, Department of Physics, University of Modena e Reggio Emilia, Modena, Italy Jennifer N. Cha • Department of Nanoengineering, UC San Diego, La Jolla, CA, USA Rahul Chhabra • University of Alberta, National Institute of Nanotechnology, Edmonton, AB, Canada Jeffrey R. Comer • Department of Physics, University of Illinois at Urbana-Champaign, Urbana, IL, USA Bernard A. Connolly • Chemical Nanoscience Laboratory, School of Chemistry, Newcastle University, Newcastle upon Tyne, UK Roberto Corradini • Dipartimento di Chimica Organica e Industriale, Univeristà di Parma, Parma, Italy Andrea Csaki • Institute of Photonic Technology (IPHT), Jena, Germany Matt V. Fagerburg • Departments of Cell Biology and Physiology and Bioengineering, University of Pittsburgh School of Medicine and Swanson School of Engineering, Petersen Institute of Nano Science and Engineering and University of Pittsburgh Cancer Institute, Pittsburgh, PA, USA Arianna Filoramo • CEA Saclay, Laboratoire d’Electronique Moléculaire, Gif-sur-Yvette Cedex, France Wolfgang Fritzsche • Institute of Photonic Technology (IPHT), Jena, Germany Alon A. Gorodetsky • Department of Chemistry, Columbia University, New York, NY, USA
- 16. x Contributors Mark M. Green • Dipartimento di Chimica Organica e Industriale, Univeristã di Parma, Parma, Italy Graham D. Hamblin • Department of Chemistry, McGill University, Montreal, Canada Ralph Hölzel • Department of Nanobiotechnology & Nanomedicine, Fraunhofer Institute for Biomedical Engineering, Branch Potsdam-Golm, Potsdam, Germany James Hone • Department of Mechanical Engineering, Columbia University, New York, NY, USA Benjamin R. Horrocks • Chemical Nanoscience Laboratory, School of Chemistry, Newcastle University, Newcastle upon Tyne, UK Andrew Houlton • Chemical Nanoscience Laboratory, School of Chemistry, Newcastle University, Newcastle upon Tyne, UK Albert M. Hung • Department of Nanoengineering, UC San Diego, La Jolla, CA, USA Yonggang Ke • Dana-Farber Cancer Institute & Harvard Medical School, Boston, MA, USA Wiepke J.A. Koopmans • Leiden Institute of Physics, Leiden Universiteit, The Netherlands Alexander Kotlyar • Department of Biochemistry, The George S. Wise Faculty of Life Sciences, Tel Aviv University, Ramat Aviv, Israel Yamuna Krishnan • Biochemistry, Biophysics and Bioinformatics, National Centre for Biological Sciences, Bangalore, India Anton Kuzyk • Lehrstuhl für Bioelektronik, Physik-Department and ZNN/WSI, Technische Universität München, Garching, Germany Christian Leiterer • Institute of Photonic Technology (IPHT), Jena, Germany Sanford H. Leuba • Departments of Cell Biology and Physiology and Bioengineering, University of Pittsburgh School of Medicine and Swanson School of Engineering, Petersen Institute of NanoScience and Engineering, University of Pittsburgh Cancer Institute, Pittsburgh, PA, USA Chenxiang Lin • Dana-Farber Cancer Institute & Wyss Institute at Harvard University, Boston, MA, USA Yan Liu • Department of Chemistry and Biochemistry, The Biodesign Institute, Arizona State University, Tempe, AZ, USA Pik Kwan Lo • Department of Chemistry, McGill University, Montreal, Canada Rosangela Marchelli • Dipartimento di Chimica Organica e Industriale, Univeristà di Parma, Parma, Italy Susan Marqusee • Department of Molecular & Cell Biology, University of California, Berkeley, CA, USA Micah J. McCauley • Department of Physics, Northeastern University, Boston, MA, USA Christopher K. McLaughlin • Department of Chemistry, McGill University, Montreal, Canada Daisuke Miyoshi • Faculty of Frontiers of Innovative Research in Science and Technology (FIRST), and Frontier Institute for Biomolecular Engineering Research (FIBER), Konan University, Kobe, Japan
- 17. xi Contributors Souvik Modi • Biochemistry, Biophysics and Bioinformatics, National Centre for Biological Sciences, Bangalore, India Khoa Nguyen • CEA Saclay, Laboratoire d’Electronique Moléculaire, Gif-sur-Yvette Cedex, France Colin Nuckolls • Department of Chemistry, Columbia University, New York, NY, USA Matteo Palma • Department of Mechanical Engineering & Applied Physics and Applied Mathematics, Columbia University, New York, NY, USA Andrew R. Pike • Chemical Nanoscience Laboratory, School of Chemistry, Newcastle University, Newcastle upon Tyne, UK Edda Reiß • Department of Nanobiotechnology & Nanomedicine, Fraunhofer Institute for Biomedical Engineering, Branch Potsdam-Golm, Potsdam, Germany Claudio Rivetti • Department of Biochemistry and Molecular Biology, University of Parma, Parma, Italy Denis Scaini • Sincrotrone Trieste, Basovizza, Trieste, Italy Stefano Sforza • Dipartimento di Chimica Organica e Industriale, Univeristà di Parma, Parma, Italy Elizabeth A. Shank • Harvard Medical School, Boston, MA, USA Jaswinder Sharma • Center for Integrated Nanotechnologies, Los Alamos National Laboratory, Los Alamos, NM, USA Michael P. Sheetz • Department of Biological Sciences, Columbia University, New York, NY, USA Friedrich C. Simmel • Physik Department, Technische Universität München, Munich, Germany Hanadi F. Sleiman • Department of Chemistry, McGill University, Montreal, Canada Thomas L. Sobey • Physik Department, Technische Universität München, Munich, Germany Naoki Sugimoto • Faculty of Frontiers of Innovative Research in Science and Technology (FIRST), and Frontier Institute for Biomolecular Engineering Research (FIBER), Konan University, Kobe, Japan Tullia Tedeschi • Dipartimento di Chimica Organica e Industriale, Università di Parma, Parma, Italy J. Jussi Toppari • Department of Physics, Nanoscience Center, University of Jyväskylä, Jyväskylä, Finland Päivi Törmä • Department of Applied Physics, School of science, Aalto University, Aalto, Finland John van Noort • Leiden Institute of Physics, Leiden Universiteit, Leiden, The Netherlands James Vesenka • Department of Chemistry and Physics, University of New England, Biddeford, ME, USA David B. Wells • Department of Physics, University of Illinois at Urbana-Champaign, Urbana, IL, USA
- 18. xii Contributors Mark C. Williams • Department of Physics, Northeastern University, Boston, MA, USA Shalom J. Wind • Department of Applied Physics and Applied Mathematics, Columbia University, New York, NY, USA Hao Yan • Department of Chemistry and Biochemistry, The Biodesign Institute, Arizona State University, Tempe, AZ, USA Hua Yang • Department of Chemistry, University of British Columbia, Vancouver, Canada
- 19. 1 Giampaolo Zuccheri and Bruno Samorì (eds.), DNA Nanotechnology: Methods and Protocols, Methods in Molecular Biology, vol. 749, DOI 10.1007/978-1-61779-142-0_1, © Springer Science+Business Media, LLC 2011 Chapter 1 Synthesis and Characterization of Self-Assembled DNA Nanostructures Chenxiang Lin, Yonggang Ke, Rahul Chhabra, Jaswinder Sharma, Yan Liu, and Hao Yan Abstract The past decade witnessed the fast evolvement of structural DNA nanotechnology, which uses DNA as blueprint and building material to construct artificial nanostructures. Using branched DNA as the main building block (also known as a “tile”) and cohesive single-stranded DNA (ssDNA) ends to designate the pairing strategy for tile–tile recognition, one can rationally design and assemble complicated nanoarchi- tectures from specifically designed DNA oligonucleotides. Objects in both two- and three-dimensions with a large variety of geometries and topologies have been built from DNA with excellent yield; this development enables the construction of DNA-based nanodevices and DNA-template directed organiza- tion of other molecular species. The construction of such nanoscale objects constitutes the basis of DNA nanotechnology. This chapter describes the protocol for the preparation of ssDNA as starting material, the self-assembly of DNA nanostructures, and some of the most commonly used methods to characterize the self-assembled DNA nanostructures. Key words: DNA nanotechnology, Self-assembly, Electrophoresis, Atomic force microscopy The notion that DNA is merely the gene encoder of living systems has been eclipsed by the successful development of DNA nano- technology. DNA is an excellent nanoconstruction material because of its inherent merits: First, the rigorous Watson-Crick base-pairing makes the hybridization between DNA strands highly predictable. Second, the structure of the B-form DNA double helix is well-understood; its diameter and helical repeat have been determined to be ~2 and ~3.4 nm (i.e., ~10.5 bases), respectively, which facilitates the modeling of even the most com- plicated DNA nanostructures. Third, DNA possesses combined 1. Introduction
- 20. 2 Lin et al. structural stiffness and flexibility. The rigid DNA double helixes can be linked by relatively flexible single-stranded DNA (ssDNA) to build stable motifs with desired geometry. Fourth, modern organic chemistry and molecular biology have created a rich tool- box to readily synthesize, modify, and replicate DNA molecules. Finally, DNA is a biocompatible material, making it suitable for the construction of multicomponent nanostructures made from hetero-biomaterials. The field of structural DNA nanotechnology began with Nadrian Seeman’s vision of combining branched DNA molecules bearing complementary sticky-ends to construct two-dimensional (2D) arrays (1) and his experimental construction of a DNA object topologically equal to a cube (2). Today, DNA self-assembly has matured with such vigor that it is currently possible to build micro- or even millimeter-sized nanoarrays with desired tile geometry and periodicity as well as any discrete 2D or 3D nano- structures we could imagine (3–8). Modified by functional groups, those DNA nanostructures can serve as scaffolds to con- trol the positioning of other molecular species (9–21), which opens opportunities to study intermolecular synergies, such as protein–protein interactions, as well as to build artificial multi- component nanomachines (22–24). Generally speaking, the creation of a novel DNA motif usu- ally requires the following steps: (1) Structural modeling: physical and/or graphic models are used to help the design of a new DNA motif; (2) Sequence design: in this step, specific sequences are assigned to all ssDNA molecules in the model; (3) Experimental synthesis of the DNA nanostructure; and (4) Characterization of the DNA nanostructure. The first two steps are crucial to pro- gram the outcome of self-assembly and assisted by computer soft- ware (25–30). In this chapter, we are going to describe the experimental protocols involved in steps 3 and 4. All chemicals are purchased from Sigma-Aldrich (St. Louis, MO) unless otherwise noted. All buffer solutions are filtered and stored at room temperature unless otherwise noted. 1. Synthetic ssDNA (Integrated DNA Techonologies, Coralville, IA) with designated sequences. 2. TBE buffer (1×): 89 mM Tris–boric acid, pH 8.0, 2 mM eth- ylenediaminetetraacetic acid disodium salt (EDTA-Na2 ). 3. 20%urea-acrylamideMix:20%acrylamide(19:1acrylamide:bis, Bio-Rad Laboratories, Hercules, CA), 8.3 M urea in 1× TBE buffer. 2. Material 2.1. Denaturing Polyacrylamide Gel Electrophoresis for the Purification of Synthetic Single- Stranded DNA
- 21. 3 Synthesis and Characterization of Self-Assembled DNA Nanostructures 4. 0% Urea-acrylamide Mix: 8.3 M Urea in 1× TBE buffer. 5. Ammonium persulfate (APS): prepare 10% water solution and store at 4ºC. 6. N,N,N,N¢-tetramethyl-ethylenediamine (TEMED, Bio-Rad). 7. Bromophenol blue (BB) or xylene cyanole FF (XC) (2×): prepare 0.1% w/v solution of the dye in 90% formamide solu- tion containing 10 mM NaOH and 1 mM Na2 EDTA. 8. Ethidium bromide: prepare 300 mL 0.1 mg/mL solution in a glass tray for gel staining. 9. Elution buffer (1×): 500 mM ammonium acetate, 10 mM magnesium acetate, 2 mM EDTA-Na2 . 10. 1-Butanol and 100% Ethanol. 11. Spin X centrifuge tube filters (Corning, Lowell, MA). 1. Polyacrylamide Gel Electrophoresis (PAGE) purified ssDNA. 2. TAE-Mg buffer (10×): 0.4 M Tris–acetic acid, pH 8.0, 125 mM magnesium acetate, 20 mM EDTA-Na2 . 1. Self-assembled DNA nanostructures. 2. 40% acrylamide (19:1 acrylamide:bis, Bio-Rad Laboratories, Hercules, CA) solution. 3. Non-denaturing loading buffer (10×): 0.2% w/v bromophe- nol blue and xylene cyanole FF in 1× TAE-Mg buffer con- taining 50% v/v glycerol. 4. DNA ladder with suitable size (Invitrogen, Carlsbad, CA). 5. TAE-Mg buffer (1×), TEMED, and 10% APS solution (vide supra). 6. Stains-All: prepare 0.01% w/v Stains-All in 45% v/v forma- mide solution. 1. Self-assembled DNA nanostructures. 2. TAE-Mg buffer (1×) (vide supra). 3. Mica discs (Ted Pella, Inc) and Atomic Force Microscope (AFM) cantilevers of choice with integrated probes (such as NP-S from Veeco, Inc for imaging in liquids). With advanced solid state synthesis chemistry, DNA synthesizer can generate DNA strands with designated sequences up to 200- base long. However, a significant yield drop is normally associ- ated with the synthesis of longer DNA strands. For example, 2.2. Self-Assembly of DNA Nanostructures 2.3. Non-denaturing PAGE for the Characterization of Self-Assembled DNA Nanostructures 2.4. Atomic Force Microscope Imaging of Self-Assembled DNA Arrays 3. Methods 3.1. Denaturing PAGE Purification of Synthetic ssDNA
- 22. 4 Lin et al. if the yield for the addition of one nucleoside is 99%, the yield for the synthesis of a 100-mer ssDNA is only ~37%. Therefore, it is very important to purify the synthetic DNA strands that are lon- ger than 30 bases to maximize the self-assembly yield in the next step. Effective ways to purify ssDNA less than 200-base long include high performance liquid chromatography (HPLC) and PAGE. Here, we discuss the protocol for denaturing PAGE puri- fication of synthetic DNA strands. 1. Set up the gel assembly following the manufacturer’s instruc- tion (we use a Hofer SE 600 Ruby from GE Healthcare) (see Note 1). 2. Mix proper volume of 20% and 0% Urea-acrylamide stock solution to prepare the acrylamide solution with desired con- centration. Each gel needs ~35 mL acrylamide solution. For example, to make an 8% polyacrylamide gel, take 14 mL of 20% Urea-acrylamide stock and mix with 21 mL of 0% Urea- acrylamide stock. Stir thoroughly to mix well. For each gel, add 262 mL of 10% APS solution and 14.7 mL of TEMED. Stir thoroughly to mix well. 3. Quickly cast the gel using 35 mL pipette and insert the comb. Make sure no air bubble is trapped in the gel. Leave the gel at room temperature for at least 30 min to allow it solidifies. 4. Prepare the DNA sample. Add DI water to each dry samples to make 0.5 OD260 /mL DNA solution. Take 4 OD of each sample (8 mL) into newly labeled tubes (see Note 2) and the rest of the samples should be stored at −20ºC. Add 2× dena- turing dye to each sample (BB, XC, or both) and add water to adjust the final volume to 20 mL. Heat the sample at 90ºC for 5 min to denature the DNA strands (see Note 3). 5. When the gel has polymerized, remove the combs and attach the upper buffer chamber (UBC) to the gel assembly. Add 1×TBE buffer (running buffer) to the UBC and rinse the wells thoroughly with glass pipette. Drain the UBC and add fresh running buffer to cover all the wells. 6. Load the samples to each well. Load 10 mL/well into the gel wells (generally 2 OD per lane) using the gel loading tips. Be careful not to flush the sample out of the well (see Note 4). 7. Carefully put the UBC and gel assembly into the lower buffer chamber (LBC) with ~3.5 L 1× TBE buffer. Add buffers into both UBC and LBC to the marked MAX lines (see Note 5). 8. Turn on the circulating water and set the temperature to 35ºC. Secure the lid of the gel box and connect the electrodes to a DC power supply. Make sure the polarity is correct. Run gel at constant current ~30–40 mA per gel for around 2–3 h depending on the length of the interested DNA fragments.
- 23. 5 Synthesis and Characterization of Self-Assembled DNA Nanostructures The tracking dye in the loading buffer provides a rough marker of the migration of DNA fragments (Table 1). 9. Turn off the power supply and circulating water. Lift the gel from the gel assembly and carefully put it into a glass try con- taining ~300 mL ethidium bromide (see Note 6). Stain the gel for 5 min and destain it for 5 min in distilled water. 10. In a dark room, lift the gel gently and put it on the UV transil- luminator. Turn on the UV at wavelength of 302 nm, use razor blade to cut the major band out (see Note 7). Turn off the UV lamp, chop the band into small pieces, and collect the small gel blocks into Spin X centrifuge tube filters. Add 500 mL of elution buffer into each filter; shake in cold room (4ºC) overnight before proceed to the next step (see Note 8). 11. Centrifuge the Spin X tube filters (4,600×g for 6 min) to separate the elution buffer from gel blocks. Add 1 mL of 1-butanol to the collected elution buffer, vortex the tube for 1 min, and centrifuge it at 600×g for 1 min. After the spin, discard the upper layer of 1-butanol with pipette into waste bottle under venting hood. The 1-butanol washing extracts ethidium bromide and tracking dyes from the DNA sample. 12. Add in 1 mL ethanol to the DNA sample and mix well. Leave the mixture in −20ºC freezer for 30 min. Spin at 16,200×g for 30 min at 4ºC to precipitate DNA. Pour out the ethanol and wash the DNA pellet with 70% v/v ice cold ethanol if desired. Centrifuge the tube at 16,200×g for 10 min after ethanol washing and pour out all liquid. 13. Use a vacuum concentrator (we use a Vacufuge from Eppendorf, Westbury, NY) to dry the purified DNA sample for 1 h at 30ºC. Add in 50 mL distilled H2 O, vortex for 1 min to dissolve the DNA sample. Measure the absorbance of the DNA solution at 260 nm (OD260 ) using a UV-Vis spectrom- eter (we employ a Biophotometer from Eppendorf) and con- vert the measured OD260 value to molar concentration using Table 1 The tracking dye migration on polyacrylamide denaturing gels (Dyes migrate to the same point as DNA strand of the indicated size in a denaturing polyacrylamide gel) Polyacrylamide concentration 5% 6% 8% 10% 12% Bromophenol blue (bp) 35 26 19 12 8 Xylene cyanole FF (bp) 130 106 76 55 26
- 24. 6 Lin et al. the extinction coefficient (e) of the DNA strand provided by the oligonucleotide vendor. Adjust the concentration of all purified DNA strand solution to 30 mM (or any other value to the experimenter’s convenience) by adding distilled H2 O. Store all DNA samples in −20ºC freezer. The formation of hydrogen bonded DNA complex is a self- assembly process. The DNA strands are mixed at stoichiometric molar ratio in a near-neutral buffer containing divalent cations (usually Mg2+ ), heated to denature and then gradually cooled to allow the ssDNA molecules to find their correct partners and adopt the most energy-favorable conformation. 1. Add stoichiometric amount of DNA strands into one 1.5 mL tube (or any other suitable tube size). Add 10× TAE-Mg buf- fer and distilled H2 O to adjust the final concentration of each DNA strand to be 1 mM or any other desired concentration. Mix well and close the tube tightly. 2. This mixture is then heated on a heat block to 95ºC for 5 min and cooled to the desired temperature by the following pro- tocol: 20 min at 65ºC, 20 min at 50ºC, 20 min at 37ºC, and if desired, 20 min at room temperature. 3. To assemble large DNA constructs, such as 2D arrays, slow annealing is desirable. In this case, the mixture is placed on a floating rack, transferred to a 2 L water bath, which is pre- heated to about 90°C and placed inside a Styrofoam box, and allowed to cool slowly to the desired temperature over the period of 2 days. This slow annealing process can also be car- ried out on a thermal cycler (see Note 9). Non-denaturing PAGE is an effective assay to characterize the self-assembled DNA supermolecules. Well-formed DNA nano- structure should migrate as a distinct band after electrophoresis. Non-denaturing PAGE also provides information regarding the yield of self-assembly. A typical gel image showing the correct formation of four helix DNA tile (31) is shown in Fig. 1. 1. Set up the gel assembly following the manufacturer’s instruc- tion as described in step 1, Subheading 3.1 (we use a Hoefer SE 600 Ruby, GE Healthcare). 2. Prepare non-denaturing acrylamide mixture from 40% acryl- amide (acrylamide:bis 19:1) stock, 10× TAE-Mg buffer and distilled H2 O. The final mixture should contain 1× TAE-Mg buffer. For example, to make an 8% non-denaturing gel, mix 7 mL of 40% acrylamide stock, 3.5 mL of 10× TAE-Mg buf- fer, and 24.5 mL H2 O. Stir thoroughly to mix well. For each gel, add 262 mL of 10% APS solution and 14.7 mL of TEMED. Stir thoroughly to mix well. 3.2. Anneal DNA Strands to Self- Assemble DNA Nanostructures 3.3. Non-denaturing PAGE for the Characterization of Self-Assembled DNA Nanostructures
- 25. 7 Synthesis and Characterization of Self-Assembled DNA Nanostructures 3. Quickly cast the gel using 35 mL pipette and insert the comb. Make sure no air bubble is trapped in the gel. Leave the gel at room temperature for at least 2 h to allow it solidify (see Note 10). 4. When the gel has polymerized, remove the combs and attach the UBC to the gel assembly. Add 1× TAE-Mg buffer (run- ning buffer) to the UBC and rinse the wells thoroughly with glass pipette. Drain the UBC and add fresh running buffer to cover all the wells. 5. Add 10× non-denaturing loading buffer to the preannealed DNA samples (finally, the DNA should be in 1× loading buf- fer). Vortex to mix well. Immediately load the DNA samples to each well using the gel loading tips. (Be careful not to flush the sample out of the well.) Take note about the sequence of the samples loaded. A DNA ladder with proper size should be loaded into a separate lane as a reference. 6. Immerse the gel assembly (together with UBC) to the 1× TAE-Mg buffer in the LBC. Add buffers into both UBC and LBC to the marked MAX lines. It is important not to disturb the sample when adding buffer to UBC. Add buffer gently along the side of the chamber. 7. Turn on the circulating water and set the temperature to 20ºC. Secure the lid of the gel box and connect the elec- trodes to a DC power supply. Make sure the polarity is cor- rect. Run gel at constant voltage ~200 V for 4–8 h depending on the size of the interested DNA complexes. Fig.1. Nondenaturing gel (8% polyacrylamide) of the 4-helix complex stained with Stains-All. Equimolar mixtures of 1 mM of each strand were annealed, and the electro- phoresis was run at room temperature. Lane M is a 100 bp DNA ladder. Lanes 1–8 contain complexes with partial combination of the component strands. Strands included in the annealing are indicated with a schematic drawing above the lane. Lane 9 corre- sponds to the full complex with all of the component strands.
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- 27. the grand staircase, where I was received by the chief eunuch, who is called kislar agaci, 'the captain of the girls.' "This giant spectre of a man ... advanced toward me, made his salaam, and ushered me, the hated, despised Giaour, into the noble marble hall of the harem, which was then for the first time polluted by the footsteps of the unbeliever. The scene around me was so singular and strange that I paused to contemplate it. The hall was of vast dimensions, supported by beautiful porphyry pillars, and the marble floor was covered with fine matting. I was now handed over to the lady superintendent of the slaves, a very wealthy woman, about twenty-four years of age, with fine dark-blue eyes, aquiline nose, large mouth, and of middle stature. "She was attired in a colored muslin dress and trousers, over which she wore a quilted lavender-colored satin paletot. Her head was covered with a small blue gauze handkerchief tied round it, and in the centre of the forehead, tucked up under it, a lovely natural dark-red rose. She wore a beautiful large spray of diamonds arranged in the form of the flower 'forget-me-not,' which hung down like three tendrils below her ear on the left side. Large diamond drops were suspended from her ears, and her fingers were covered with numerous rings, the most brilliant of which were a large rose-pink diamond and a beautiful sapphire. Her feet were encased in white cotton stockings, and patent-leather Parisian shoes. Her name was Anina: she had been formerly an Ikbal 'favorite.' ... The lady superintendent now took me by the hand, led me up two flights of stairs covered with thick, rich Brussels carpet of a most costly description, and as soft and brilliant in colors as the dewy moss of Virginia Water. The walls were plain. Then we passed through a suite of several rooms, elegantly carpeted, in all of which stood long divans; some of which were covered, with white, and others with yellow and crimson satin. Over the doorways hung white satin damask curtains, looped up with silk cords and
- 28. tassels to correspond, with richly gilded cornices over each. ... Against the walls were fixed numerous silver chandeliers, each containing six wax candles, with frosted colored glass shades made in the form of tulips over them. On each side of the room large mirrors were fixed in the wall, each of which rested on a marble-topped console table supported by gilded legs. The only other articles of furniture that were scattered about the apartments were a dozen common English cane-bottom Kursi- chairs." She is next conducted further on to some dormitories, where bedsteads are wanting, being an article of furniture unused by the Gypsies. Against the walls were piled up beds in heaps, covered over with a red silk coverlet. On the divan was placed a silver tray —both toilet-tables and wash-hand-stands being unheard-of comforts—containing the princesses' toilet requisites. In her general inspection the governess is led to the apartments of the Princess Epouse, the mother of the little boy for whom Mrs. Lott is engaged. This princess is dressed—but let dame Emeline describe the scene, as only a lady can do it: "The Princess Epouse, attired in a dirty, crumpled, light-colored muslin dress and trousers, sat à la Turque, doubled up like a clasp-knife, without shoes or stockings, smoking a cigarette. ... Her feet were encased in babouches, 'slippers without heels.' ... In front of the divan, behind and on each side of me, stood a bevy of the ladies of the harem, assuredly not the types of Tom Moore's 'Peris of the East,' as described in such glowing colors in his far-famed Lalla Rookh, for I failed to discover the slightest trace of loveliness in any of them. On the contrary, most of their countenances were pale as ashes, exceedingly disagreeable, flat and globular in figure; in short, so rotund, that they gave me the idea of large full moons; nearly all were passé. Their photographs were as hideous and hag-like as the witches in the opening scene in Macbeth, which is not to be wondered at, as some of them had been the favorites of
- 29. Ibrahim Pasha. ... Some wore white linen dresses and trousers. Their hair and finger-nails were dyed with henna. ... They had handsome gold watches ... suspended from their necks by thick, massive gold chains. Their fingers were covered with a profusion of diamond, emerald, and ruby rings; in their ears were ear-rings of various precious stones, all set in the old antique style of silver. ... Behind stood half-a-dozen of white slaves, chiefly Circassians." The mother leaves a favorable impression on the mind of the governess, who, being finally dismissed from the interview, pursues her explorations and makes a great discovery neither complimentary to the princess nor cleanly, where water is abundant, but where ablutions seem to be abnormal; for it is written in her journal that "Thence we passed along a stone passage which leads to her highness's bath-room. ... The marble bath is both long and wide, with taps for hot and cold water. The water actually boils into which their highnesses enter. This only occurs when they have visited the viceroy, and not daily, or even at any other time. The bath of the poets is a myth." The governess at last reaches her own chamber, where she is destined to sleep and seclude herself in her leisure hours. The prospect at first is not inviting, nor does a second view afford more encouragement; an evident sense of disappointment, if not of dismay, is experienced; and thus she pours forth her vexation: "On the right-hand side of the first room was the small bed- room which was assigned to me as my apartment. It was carpeted, having a divan covered with green and red striped worsted damask, which stood underneath the window, which commanded a fine coup-d'ceil of the gardens attached to the palace of the viceroy's pavilion. The hangings of the double doors and windows were of the same material. The furniture
- 30. consisted of a plain green painted iron bedstead, the bars of which had never been fastened, and pieces of wood, like the handles of brooms, and an iron bar, were placed across to support the two thin cotton mattresses laid upon it. There were neither pillows, bolsters, nor bed linen, but as substitutes were placed three thin flat cushions; not a blanket, but two old worn- out wadded coverlets lay upon the bed. Not the sign of a dressing-table or a chair of any description, and a total absence of all the appendages necessary for a lady's bed-room; not even —" Well, well, Mrs. Lott, the "not even" was, in your civilized opinion, certainly very odd to be sure. But don't mind trifles; let it be forgotten; let us ramble elsewhere. You were saying just now something about four broad steps; go on; that's right. "Four broad steps led down into the garden, close to a plain white marble-columned gate, on the top of which stood out in bold relief the statues of two huge life-sized lions. ... Here and there were scattered rose-trees, the brilliancy of whose variegated colors and the perfumes of their flowers were delightfully refreshing; geraniums of almost every hue; jessamines, whose large white and yellow blossoms were thrice the size of those of England, and a variety of indigenous and eastern plants, shrubs, and flowers, which were so thickly studded about that they rendered the view extremely picturesque, and perfumed the air, grateful to the senses. Verbena trees, as large as ordinary fruit-trees; other plants bearing large yellow flowers, as big as tea-cups, with most curious leaves; cactuses, and a complete galaxy of botanical curiosities, whose names the genius of a Paxton would be perhaps puzzled to disclose, ornamented those Elysian grounds." This is only one sketch of only one spot in the many gorgeous and luxurious localities. Space forbids copying more; but the book
- 31. states: "Leaving these neglected scenes of amusement, we proceed along a path to the right, through a superb marble-paved hall, the ceiling of which is in fresco and gold. It is supported by twenty-eight plain pink-colored marble columns, surmounted by richly-gilded Indian wheat, the leaves of which hang down most gracefully, on each side of which, and also above ... are some very handsome lofty rooms, the ceilings of which are also in fresco, with superb gilded panels. ... "The grounds of Frogmore, the Crystal Palace, St. Cloud, Versailles, the Duke of Devonshire's far-famed Chatsworth, and our national pride, Kensington Gardens and Windsor Home Park, exquisite, beautiful, and rural as they are ... all lack the brilliant display of exotics which thrive here in such luxuriance. The groves of orange-trees, the myrtle hedges, the beautiful sheets of water, the spotless marble kiosks, the artistic statuary, are all so masterly blended together with such exquisite taste, that these gardens ... completely outvie them." The princesses were sometimes as highly adorned as the halls of marbles and frescoes, and as ornamental as the gardens of blooming exotics. On the festival of the Great Bairam, or on state occasions, when lady visitors made formal calls to compare complexions and cashmeres, their highnesses are spoken of with the highest delight: "They wore the most costly silks, richest satins, and softest velvets; adorned themselves with the treasures of their jewel caskets, so that their persons were one blaze of precious stones. That crescent of females (for they always ranged themselves in the form of the Turkish symbol) was then a parterre of diamonds, amethysts, topazes, turquoises, chrysoberyls, sapphires, jaspers, opals, agates, emeralds, corals, rich carbuncles, and rubies. In short, the profusion of diamonds
- 32. with which the latter adorned their persons from day to day became so sickening to me that my eyes were weary at the sight of those magnificent baubles, to which all women are so passionately attached." But weary as were her British eyes, still she gazed in rapture when the darling gems were on exhibition; moreover, in the journal the impressions were faithfully recorded. On another occasion, when some princesses were coming, "The Princess Epouse, the mother of my prince, was attired in a rich, blue-figured silk robe, trimmed with white lace and silver thread, with a long train; full trousers of the same material, high-heeled embroidered satin shoes to match the dress. On her head she had a small white crape handkerchief, elegantly embroidered with blue silk and silver, and round it placed a tiara of May blossoms in diamonds. She wore a necklace to correspond, having large sapphire drops hanging down the neck. Her arms were ornamented with three bracelets, composed of diamonds and sapphires, and an amulet entirely of sapphires of almost priceless value. ... At times my eyes, when looking at the Peris arrayed in all their gems, have become as dim as if I had been fixing them on the noonday sun." What young lady of an enterprising turn of mind would not be willing, after reading these glowing descriptions, to pack up her Saratoga trunks, to engage the Adams Express Company, and to charter the Cunard line of steamers, to aid her on to a glorious future near the base of the pyramids? Certainly not one of the ambitious and strong-minded. But they need not ask the English governess to go with them. She has been there; she will respectfully decline going again—not she, as Shakespeare's other old lady in Henry the VIII. exclaims, "not for all the mud in Egypt." For another part of the story remains to be told; another side of the picture to be presented; and dame Emeline tells it truthfully,
- 33. she paints it lifelike; the rose is beautiful, but beware the serpent under it. Mrs. Lott is apparently a gentlewoman, refined, accomplished, intellectual, with an appreciation of the difference between civilized society and barbarism. But in the vice-regal harem, education was not to be found; ignorance was universal, superstition reigned supreme. None could read, or write, or sketch, or converse on a rational subject. No one could sing or perform on a musical instrument; none cared for to-morrow or for a hereafter. Their daily routine had all the monotony of the desert with its burning sands, destitute of variety in incident or shade of change; it was equally unproductive and utterly worthless. They had nothing to expect with pleasing anticipation; they had nothing to remember with delight. Physically, morally, mentally they were unclean and debased. Their passions, when aroused, were ungovernable; their greatest joy was revenge upon a rival; and their revenge was deadly, by suffocation or submersion, poison or the bow-string. Their amusements were all sensual; their weary hours of listless idleness were passed in indulgence of some enervating vice alike deleterious to health, comfort, and color. The servants were steeped in only a lower depth of dirt and depravity. The princesses had the power of life and death over them, and it was a power often exercised; they would put them to the torture for a trivial fault, the breaking of a plate or the falling of a cup; and cheeks and arms seamed with parallel rows of the red- hot iron, attested how often and how unmercifully cruel had been their punishment. The food of the menials was not prepared for them, nor given to them; but they purloined by stealth from the dishes on their way to the princesses' apartments; and after their repast was ended, the refuse of chicken and pigeon bones, of mutton, of soup, of rice, of vegetables, and the rinds of fruit were tossed into a basket in one loathing mess, mixed up, around which the servants flocked like carrion birds, and, squatting on the floor,
- 34. inserted ravenously their reeking hands to pick out disgusting morsels with their dripping, unwashed fingers. The laundry did not require much water; for the volume informs us, "Those who performed the duties of washerwomen were occupied daily in their avocation, except on the Sabbath, (Fridays.) But that was not very laborious work, since neither bed, table, nor chamber linen are used. Thus they were engaged until twelve, when their highnesses partook of their breakfast separately. It was served up on a large green-lackered tray, minus table-cloth, knives and forks, but with a large ivory tablespoon, having a handsome coral handle, the evident emblem of their rank as princesses. It was placed upon the soofra, a low kind of stool, covered with a handsome silk cloth. The repast occupied about twenty minutes. Then pipes, in which are placed small pills of opium, or more often cigarettes and coffee, were handed to them, and each princess retired to her own apartment. Thus they became confirmed opium- smokers, which produced a kind of intoxication." ... Their common indulgence in opium, with a profuse supply of European wines and Schiedam gin, produced its natural results, and is thus depicted: "Oftentimes after the princesses had been indulging too freely in that habit to which they had became slaves, their countenances would assume most hideous aspects; their eyes glared, their eyebrows were knit closely together; no one dared to approach them. In fact, they had all the appearance of mad creatures, while at other times they were gay and cheerful. "They only combed their hair (which was full of vermin) once a week, on Thursdays, the eve of their Sabbath, (Friday, Djouma;) when it was well combed with a large small-tooth comb; and pardon me, but 'murder will out,' the members of the vermin family
- 35. which were removed from it were legion. It was afterward well brushed with a hard hair-brush, well damped with strong perfumed water. Their highnesses never wore stockings in the morning, nor did they change any of their attire till afternoon." When the summer heats set in, the harem was transferred to the coast at Alexandria, to inhale the fresh breezes from the sea. The preparation for flight was attended with some rich scenes and ludicrous exhibitions. But their transit on the railroad, boxed up like pigs or poultry on a cattle-train, is indescribable in a decent print. The prelude to the trip will bear repeating; it is an amusing contrast with the festal robes on the day of the Great Bairam; the cutaneous sensation it excites is the penalty to pay for the knowledge imparted; the company is right regal. "As soon as orders had been given to the grand eunuch to hasten the departure of the vice-regal family to Alexandria, ... there was bustle all day long. One morning when I returned from the gardens, ... I entered the grand pasha's reception- room; ... there were their highnesses, the princesses, squatted on the carpet amidst a whole pile of trunks. They were all attired in filthy, dirty, crumpled muslins, shoeless and stockingless; their trousers were tucked up above their knees, the sleeves of their paletots pinned up above their elbows, their hair hanging loose above their shoulders, as rough as a badger's back, totally uncombed, without nets or handkerchiefs, but, pardon me, literally swarming with vermin! No Russian peasants could possibly have been more infested with live animals. In short, their tout ensemble was even more untidy than that of washerwomen at their tubs; nay, almost akin to Billingsgate fisherwomen at home; for their conversation in their own vernacular was equally as low. They all swore in Arabic at the slaves most lustily, banged them about right and left with any missile, whether light or heavy, which came within their reach."
- 36. At last the governess lost her health. The food was too unsuitable for a Christian woman, and the atmosphere, redolent of the overpowering rich perfumes of the gardens mingled with sickening, stupefying opium smell and smoke, along with other odors, almost intolerable. After visiting Constantinople with the harem, she threw up her engagement and returned to England. This abasement of woman is not to be wondered at; for wherever the Christian idea of marriage is lost or subverted, woman becomes the mere object of passion, and degradation is sure to follow.
- 37. Translated From Etudes Religieuses, Etc., Par Des Peres De La Compagnie De Jesus. The Flight Of Spiders. A Paper Read Before The French Academy Of Science, March, 1867. About fifteen years ago, I was sitting in an arbor of my garden, reading, when a little spider fell on my book, whence I could not tell, and commenced to run over the very line I was reading. I blew hard to chase him away, but he would not go. He lifted himself strangely up, and I cannot explain how, but he lodged on a sprig of verdure just above my head. "Well," said I, "for a little animal like that, this is a wonderful feat! How has he accomplished it?" To satisfy myself, I took him up again, balanced him on my book, and, after assuring myself that he had no invisible thread to aid him, I blew again, and again the little fellow did the very same thing. With redoubled curiosity, I tried him once more, and, to see better, I sat down in the bright sunlight. Again I balanced him on the book, looked at him as closely as possible, and, when I felt assured no precaution could have escaped me, I blew once more. ... Resuming the same inclined position, the spider as quick as lightning darted the finest possible thread out of him, raised himself in the air, and disappeared. I confess I was stupefied. Never had I imagined these little animals could fly without wings; so I consulted several works on zoology, but I was astonished to find there was no mention made of the flight of spiders, nor of the ejaculatory movement of which I had witnessed so curious an example. [Footnote 96]
- 38. [Footnote 96: In M. Eugène Simon's Natural History of Spiders, the most recent work of the kind, he says, speaking of the manner in which l'épéire diadème constructs its web: "Several authors suppose that the spider darts its thread like an arrow, others imagine it throws it upward in the air while flying as a fly would; but neither of these explanations rests on observation, and they are, after all, simple hypotheses." Then, describing his own observation as to how a spider acts to make fast its great threads, he says, "It seems to take a horizontal position, and moves contrary to the wind." M. Simon's work gives us nothing else to lead us to suppose he has observed the wonders spoken of.—Tr.] So there was a new question presented to me, and my vocation to study the habits of these little animals—which hitherto had given me no concern—decided for me. I immediately lost all repugnance, all distaste, and threw away all the unjust precautions of which the spider is too often the object, and of which I was as culpable as any one else. And from that time I welcomed its appearance; was most happy to meet with it, looked for it, indeed, and studied its habits almost with furor. And I can say that, thanks to this hearty preoccupation, which never left me, I found every opportunity to follow my inclination, and knew where to find spiders in all sorts of unheard-of places. Such are the singular effects of curiosity once excited, and still another proof that, in order to study nature well, we need only a mysterious glimpse of the unknown to redouble all our energies to explain it thoroughly. And as in this study, trifling as it may appear, I seem to have met with facts not known hitherto, but which deserve to be understood, I here resume the principal ones: those that treat of the flying of spiders; of the habitation of some species in the air; and of the gossamer or air threads—a singular phenomenon, for a long time
- 39. discussed in vain, but which I believe I have definitively solved. I only ask the naturalists to judge one fairly, not by theory, but by facts. And I am persuaded, if they will take the pains to verify what I advance, they will find me exact; and, if they begin doubtingly, I hope, after they have read my observations, they will conclude as others to whom I have communicated them. Mocking and incredulous at first, they have ended by believing their own eyes, and testifying to the evidence presented to them. May my labor prove useful, and, above all, contribute to the glory of the great God, whose just title is, Magnus in magnis, maximus in minimis. I. Threads Thrown Out By Spiders. The first thing that I perceived, and that put me on the track of the rest, was, as I have just said, that the greater part of aranéides, especially certain varieties of thomises lycoses, etc., besides the thread that they always draw with them, have the power of darting one or more of extraordinary length, and of which they make use to accomplish distances, to fasten their webs from one point to another, and even, as we shall see further on, to raise themselves in the air and there to seek their prey. The spider always points his abdomen to the side where he wishes to go. The thread shoots like an arrow, fastens itself by the end to the place destined, and the spider passes as under a suspended bridge. If this thread is cut, it is immediately replaced by another; and the ejaculation is so prompt, so rapid, the thread so straight, so tenuous, so brilliant, that it might be taken, if I may so express myself, for the jet of an imperceptible ray of light. To perceive this clearly, the spider must be held on a level with the eyes, which should be shaded, and examined with one's back to the sun. The best time for such an observation is in the morning or evening, when the sun is low in the horizon and the temperature is mild; for
- 40. without this latter condition the torpid spider is more inclined to creep along the earth than to throw out new threads. Sometimes, to excite them, they may be held by their ordinary thread and gently shaken or blown upon—just a few puffs of breath —which they detest. I have thus been able to scan closely, while watching their development, this instantaneous jet of thread, which could not be less than five or six yards long, that is, fifteen hundred or two thousand times the length of the spider. What a tremendous apparatus must be necessary to these little animals for so rapid an ejaculation, and one so disproportioned to their size! And especially if we consider that this thread, inasmuch as it adheres to the animal, has not the appearance of an independent organ, but seems solely to obey its will. Thus I have seen spiders, who seemed to miss the end desired with the first stroke, continue to hold the thread in the same direction, and actually palpitate, if I may so say, while striving to make it adhere. But a truly interesting sight, and one obtained at a very trifling expense, is that which the thomises bufo offer, described by Walckenaer, in the first volume of his History of Insects, page 506. In truth, these araneides do not only throw out one thread, but an entire bundle of them, and are seemingly guided by the smaller threads, just as a peacock unfolds by degrees his splendid plumage. And even in one's own room this sight may be enjoyed. It is only necessary to collect these thomises and keep them in separate boxes, and nourish them in winter with one fly or so a month. Then take the boxes out, put them on a table in a very warm room, and sit a little in the shade and watch them. Very soon from each box will appear a multitude of threads, of extreme freshness and fineness, which the spider throws into the air with
- 41. inexhaustible profusion. At certain seasons of the year we can enjoy this spectacle again, and at even less expense. II. Flight Of Spiders. Another property not less remarkable that these araneides possess (thomises bufo, lycoces voraces, etc.) is that of flying; that is to say, of elevating themselves in the air, there sustaining themselves, and travelling about horizontally and vertically, with or without a thread; in a word, acting exactly as if in their own element. This fact I have witnessed a thousand times, and it has been certified to by a great number of people, who, at first incredulous, and alarmed for the laws of gravitation, were compelled to confess the reiterated testimony of their own eyes. I had some pupils under my charge, and to them this study became a continued source of amusement. During their recreation, they found suitable spiders for me, and, when they brought them to me, I rested them on my fingers and made them mount upward in the air; and invariably, after having watched them for some moments, they were entirely lost to sight. But when I made the discovery—of which I will speak later—of the general migration which some species make yearly toward certain regions of the atmosphere, I had no longer any trouble to enjoy this performance to my heart's content. The flight of spiders is sometimes very rapid, particularly when they start. They often escape from one's hands while they are carefully watched. This happened to me one day with a voracious lycose that I had for a long time importuned without success. Just as I was going to give him up as entirely stupefied, he suddenly escaped from me by a lateral movement, so rapid that for a moment I lost sight of him; but, when I found him a moment afterward, he was suspended quietly in the air. I also remarked that he set out without throwing any thread, and this was not the only
- 42. time I made the same observation. I was experimenting one day with some amateurs in the interior court of the college where I live, and, having started a lycose, we saw him occupy himself at first with the neighboring galleries, running up and down for about twenty yards, about a tenth of a yard from the arch, against which he knocked himself from time to time, and groped about to look for a passage; not finding one, he threw himself back into the court, raised perpendicularly, and disappeared toward the clouds. His thread, if he had one, could not have been longer than a tenth of a yard. Ordinarily, however, before they ascend, they throw out a thread which they follow for a short time; then, arriving at a certain height, they break it, in order to navigate more easily. If any is left before them, they wind it rapidly with their feet, throw it aside, and form those pretty little crowns of white silk in form of cracknels, that we often see flying in the air in time of gossamers. Again, they balance themselves quietly with a thread which rises perpendicularly above them, and gives them the appearance of floating. But a peculiarity still more remarkable in the flight of spiders is the attitude that they take in flying. They generally swim backward, that is to say, the back turned from the earth, the feet folded on the corselet, and perfectly immovable. How can such a flight be explained, for they are already heavier than the air? Plunged into alcohol, they sink quickly; but in the air they seem to possess an ease, a liberty, a facility of transport, so admirable that I have never been able to see in them the slightest motion, nor even an apparent increase of weight. Does not this fact present an interesting question for the skilful to contemplate? III. How Long They Can Remain In The Atmosphere? At this portion of my history I have to relate facts the most curious and unexpected; and, unfortunately for me, more true than probable. I acknowledge I was loath to publish them, or assume
- 43. concerning them any responsibility. But I was firmly convinced, and therefore hoped to be believed, especially by this generation of fearless naturalists, who are astonished at nothing in nature, and who, having often been surprised in the relation of almost incredible marvels, must certainly make allowances for a few more in another quarter. Let us look at, for instance, the wonderful things related of the argyronete, or aquatic spider. [Footnote 97] [Footnote 97: The argyronete is a spider that lives in the water where she constructs a charming little edifice that appears surrounded with a silky mortar. The down that covers her contains a certain quantity of air for respiration. This gives her in swimming the appearance of a ball of quicksilver, from which we have her name.] I could not tell anything more unlikely, so I will only exact for the atmosphere a companion to what the Père de Lignac discovered in the last century for the water. Yes, I pretend there are spiders that live in the air as well as those living in water, and that every year, from the earliest days of spring, there is, unknown to us, a general migration of spiders toward the atmosphere, where they pass their best season, form their nets, chase their prey, and only return to earth in the first fogs of autumn to find their quarters for the winter. I add, also, that this ascent and descent give rise to the curious phenomenon, still so badly explained, of the gossamer. And as it was to the study of this phenomenon that I owe my knowledge of the rest, may I be permitted here, by way of demonstration, to relate briefly the path I have followed and the proofs which have led to the conviction I express? Attracted, as I was, by all that concerns spiders, I could not remain indifferent to a fact so important and interesting as the periodical apparition of those threads which in spring and autumn we see flying about in long white skeins, clinging to trees, to hedges, and
- 44. to the vestments of the passers-by, carpeting the country in a few hours with more silk, and finer and whiter, than could be spun in a year by all the reels in the world. Admirable netting, glistening in the light of the setting sun, and reflecting the sweetest, softest tints of gold, vermilion, and emerald, and receiving the pretty and poetical name of "fils de la Vierge." Was there not between this phenomenon and my preceding observations a secret tie, some mysterious relation? I seemed to foresee it, and, setting to work immediately, rejected from the very beginning the usual explanation of this phenomenon. How, indeed, can we admit these floating gossamers as merely the refuge webs of spiders, torn by the violence of the wind from the trees and forests and carried capriciously through the air? Will not the slightest observation convince us that they never appear but in the calmest moments, on days foggy in the morning, but afterward beautiful, and not preceding a storm; never in summer, often in the spring and autumn, and sometimes even in winter? If the winds carry them, why do they not appear in summer? Are violent winds and spider-webs both wanting? And who has ever seen one of these webs carried by a hurricane, especially in quantity sufficient to produce such a phenomenon? For the fall of gossamers sometimes lasts for almost entire days, and in certain countries during the middle of the day the fields are covered with them. Add, too, that violent winds are generally local, while this phenomenon is universal, and so periodical that in the same climates it appears at the same epochs, and, when one knows what produces it, it is easy to predict the time and day of the apparition. Discontented, then, on this point with books and their explanations, I turn completely to the side of nature, and present all I observed. From the first appearance of these threads in autumn, I was struck with the immense multitudes of new spiders met with everywhere, and which I had not seen during the summer. Little brown lycoses filled the air, so that it seemed as if it had rained them. If one
- 45. walked in the fields, the meadows, the gardens, on the borders of the woods, among heaps of dried leaves, scattered all through the forest everywhere, could be seen myriads of these little brown spiders, jumping up and flying before me in every direction, and exactly such as I had already recognized as such excellent swimmers. After having passed the winter in the earth, in the holes of worms that they completed with a little silk, they reappeared after the cold in great numbers, to disappear again entirely in the first bright days of spring, and as if by enchantment. If one is seen again during the summer, we may be sure it is some female retarded by laying her eggs, and dragging laboriously her cocoon after her. Now, what has become of the others? For several months I could not satisfy myself on this point, when, on the 21 St of October, 1856, in the enclosure of the little seminary of Iseure, near Moulins, I came to a positive decision, I was observing the fall of a large quantity of gossamers, which were falling on that day in large white flakes, when I perceived close to me in the air one of those little black spiders descending gradually, and as if she were jumping. She held by an invisible thread to a large flake, which came down slowly about seven or eight yards above her; but, keeping outside of it, she hung by the end of the long thread, like an aeronaut underneath his balloon. My attention once attracted, I noticed so great a number that I was astonished I had not taken care sooner; for there was scarcely a flake underneath which there were not one or two, and this sometimes even before the flake itself was visible. [Footnote 98] [Footnote 98: There is an observation which confirms my own. We read in Darwin's Journal, page 159: "Mr. Darwin saw a large number of gossamers on the ship Beagle, when she was about 60 miles from the mouth of the Rio de la Plata. It was the first of November, and these gossamers were carried by a very light breeze, and on each were found an immense number of little spiders, similar in appearance, about the twelfth of an
- 46. inch in length, and in color a deep brown. The smallest were a deeper shade than the others. None were found on the white tufts, but all on threads." Journal of Researches into the Natural History and Geology of the Countries visited during the Voyage of his Majesty's Ship, the Beagle, 1845.] Each one was separated by a slender thread, and followed the motion of its balloon. If they met a tree or a bush, they landed upon it; if not, coming close to the earth, they ran along and were lost in the verdure. If I approached them too quickly or made a noise, they remounted rapidly by their threads and went to disembark somewhere else. I also examined some of the flakes. They were all shining white mats, appearing as if they had been washed. Several contained wings and feet of flies, fragments of the case of little coleoptera, and other remnants of their aerial festivities. This encounter was for me a revelation. I knew where the spiders, whom I had seen disappear so brusquely, took refuge, and, however rash my judgment may appear, I felt assured I had solved an interesting problem. But to establish seriously and give to science an opinion so new and original as that the atmosphere may be peopled with spiders, I soon felt that more proof was necessary in order to sit down calmly under my personal conviction. So I concluded I should not be doing too much if I added to the verification of their descent that of their ascension, and could surprise them in this new migration. I waited, therefore, impatiently for the spring. But that spring, and for five or six that followed it, great was my disappointment; for, though I perceived several isolated ascensions, yet nothing in the proportion I had imagined or that could justify my hypothesis. I began then to doubt seriously my success, when
- 47. an incident occurred that relieved my embarrassment, and proved how trifling sometimes are the causes which lift the veil from nature. I was looking straight upward, but sitting close to the earth, and so as to be able as much as possible to exclude the sun from my eyes. And here, by the way, a fact is made palpable, by no means microscopic, but which has escaped so long not merely the observation of the crowd of vulgar observers, but of those even who are wide awake and study carefully; namely, that it is not necessary to carry one's nose always in the air, if I may so express myself, to examine closely, to investigate, or to render a faithful account of phenomena. On looking upward—as an ascension only takes place on very beautiful days, succeeding generally to bad weather—spiders cannot be distinguished from the multitude of other insects which fill the air. But if, on a beautiful day, mild, calm, and brilliant in sunlight, succeeding as nearly as possible to a rain warm with the south wind, at about nine or ten o'clock in the morning, a post is chosen on an eminence of a meadow or an avenue, and there, as near the ground as may be, and crouching low, the observer will look horizontally, he will perceive a series of fire-works, formed of innumerable threads launched from every direction and inclined toward the sky. This is the prelude. Soon the spiders detach themselves and mount slowly by their threads. The most conspicuous are the thomises bufo, because they are the largest, and because they only ascend with an entire bundle of threads, which gives them the appearance of small comets. Thus have I decided 1st. That there is not only one ascension every year, but several, at least partial ones; that they do not always take place in spring, but often in the autumn, and sometimes even in the winter; and in general, from the descent which has taken place in the beginning of autumn until the definitive ascension in the spring, there are but few favorable days of which the spiders do not profit to make an
- 48. aerial journey, or at least to throw out a large number of threads. Thus, in the Beaujolais, where I have lived for several years, there were partial ascensions on the 1st, the 19th, and the 28th of November, 1864; the 21st, the 23d, and especially the 25th of October, the 9th of November, and the 6th of December, 1865. In 1866, the 18th and the 30th of January, the 3d of February, the 3d, 14th, and 31st of October, and the 17th of December. In 1867, the 10th of February, ... the last, however, less considerable than might have been predicted by the beauty of the day. The day previous was so mild, though cloudy, that many of the spiders may have embarked incognito. Many, also, may not have judged it a propos to fly away, for a great number still remained on the ground. I forgot to observe the temperature of all the days I have noted. The director of the Normal School of Villefranche having had the kindness to show me the meteorological register which he had kept with great care, I was able to prove that in calm weather only ten or twelve degrees of heat were necessary to induce them to mount upward. The least exposed begin; then immediately the others, so soon as the heat reaches them; but after three or four o'clock in the afternoon no more ascensions are perceived, unless they are provoked; and this does not always succeed, 2d. Before taking their flight, they generally cling to some elevated object that they meet with easily, such as shrubs, bushes, props of vines, or blades of grass escaped from the scythe. To these they affix their threads and warm themselves well in the sun before commencing their excursion. This is the happy moment for amateurs to make their observations, for there is scarcely a blade of grass that does not contain one or more; and, if the branches of young trees are suddenly struck with a slight blow, a great number are detached, suspended at the end of their threads; and very often rare specimens are thus found not discoverable elsewhere. IV. To What Height Do They Raise Themselves in the Atmosphere?
- 49. On this point I have not been able to make any direct observation. Perhaps I have dreamed of offering objections to the concourse of intrepid human navigators who undertake such perilous excursions in the air, and for my interest in the study I have found two excellent reasons. The first, that it would be well for them to know that, if they have not had rivals, they have had precursors, who, for 6000 years, have executed silently and noiselessly what they have claimed for themselves by every effort of puffs and publicity. The second, and a still more serious objection and that I believe will truly interest the future in this young industry, is that if the argyronete and its bell has given to science the instrument with which the divers explore the depths of the sea, why may not the study of aerial spiders furnish for aeronauts—these divers in air— the complete apparatus which they require to raise themselves to any height, direct their movements, and maintain themselves at will? Have not these little animals resolved this problem for centuries? Yet the present state of aerostation does not afford ground sufficient for comparison. We are, therefore, reduced to conjecture; and, if I may be permitted to express mine, this is what I think: I believe that spiders rise to the same height where on the fine days of summer one can see the swallows and martins hover, almost lost to sight, in pursuit of gnats that people these regions of the atmosphere. I found this belief on the webs of spiders seen falling in autumn, that seem to come at least from nearly such heights. They begin to be seen at a hundred or a hundred and fifty yards, and there is no great temerity in affirming that they have already traversed a good part of their course. An observation made in 1864, if conclusive, would tend to make remoter still the habitation of spiders; for the fog that determined the fall that year was a high fog, that is to say, one of those uniform mists that hide the sky for several days together, and seem to extend to a great height. But, I repeat, this is all conjecture. One good observation would have been worth far more.'
- 50. V. Conjectures On The Mode Of Building Of Spiders In The Air. Perhaps here I should stop, and, having stated facts, leave to others their explanation. How do spiders sustain themselves in the air? How can they so long brave the winds, the rains, the storms; arrange their webs in emptiness and without apparent means of support? Prudence counsels me to avoid these questions, but my rôle of simple observer permits them. However, in waiting for better things, I decide still to hazard some conjectures, were it only to prove that a fact once admitted, it would not be absolutely impossible for the wisest to explain it. The first idea that came to me was that these spider-webs raise themselves in the air as the kites of children, and, made fast to the tops of trees and edifices by long threads, they are sustained by their own lightness. This idea was suggested to me by a sight I was witness to one day at the Seminary of Vals, near Le Puy. From a corner where I was in shadow, I perceived distinctly on each high ridge of the roof, lightened by the rays of the sun, long threads which rose perpendicularly in the air, like large cords, balancing themselves slowly right and left, without ever going out of a certain field of oscillation. But I soon gave up this idea. How admit, in truth, that on two or three threads, and without any other means of support, spiders could weave their true webs? Would not some of these aerial constructions tumble down every day, ruined by their own weight? while it is acknowledged they only fall in autumn, and always together. I therefore rather incline to believe that the spiders are sustained in the air by the distention of an interior vesicle, analogous to that of fish, and that they ejaculate by their threads, which are numerous, and pierced with an infinity of little tubes, large bundles of threads, by which are taken the insects that serve for their prey; that they resist the winds as fish do the tossing of the sea, and their threads,
- 51. being glutinous, are not dampened by the rain; and also being excellent conductors of caloric, as is proved by the abundant drops of dew which they pearl near the earth, on the hedges, etc.; and if after a calm night they are touched by an autumn fog, these heavy and moistened threads weaken and fall one over the other, and form the silky flakes that are seen from ten to eleven o'clock in the morning, flying about in cloudy days with the spiders who inhabited them during the summer. This, hoping for better, is the explanation I hazard, and I submit it with the rest to the appreciation of competent men. If only these pages attract attention to a merited subject, and provoke numerous observations, which alone can ever fully elucidate it, the author will be more than repaid for the few researches he has presented in this article.
- 52. Translated From The "Revue Du Monde Catholique." John Tauler. By Ernest Hello. History has an astonishing memory. She records the day and hour of battles with exact fidelity. She knows a thousand things. She has recently discovered, if I do not mistake, the name of Julian the Apostate's cook. She remembers everything of little importance. The names of celebrated mistresses who have amused or poisoned renowned personages, are transmitted from age to age. Erudition has been making strides during the last hundred years, as if she had seven-leagued boots. To deserve the admiration and gratitude of mankind, however, she should not have degraded herself, but taken a higher sphere in her progress. Her memory indicates greatness of genius; but she is like calumny, she increases in size as she advances through the centuries. In her labors, researches, and exploits, she has been mostly busied with soldiers, and frequently forgotten God and man. She could not think of everything at once; the hidden history of humanity is yet to be written; the greatest events of the world are secret to this very day; and those who reflect on them are men of a special caste. If there were question of the battle of Marathon, or of Antony and Cleopatra, our contemporaries would be found well instructed; but do they know John Tauler, the German Tauler, of the Dominican or preaching order? Master Tauler was a great preacher—powerful and popular. One day he gave a learned discourse, in which he taught the way of perfection, with all his characteristic assurance. To become perfect, he enumerated twenty-four conditions, which he developed before an attentive and brilliant audience. After the sermon, a layman, one
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