Next Generation Sequencing
- 1. Seminar on : Next Generation Sequencing Presented by, Miss AMRITHA S R BSc BIOTECHNOLOGY FACULTY OF LIFE SCIENCES JSS ACADEMY OF HIGHER EDUCATION AND RESEARCH 1 Dated on : 1st March 2019
- 2. Overview Genome History NGS Procedure ( Ion torrent , 454) Application 2
- 3. GENOME Genome is the sum total of genetic material . The amount of DNA present in a genome of a species is called C-value. In lower eukaryotic organisms like yeast, amount of DNA increases with increasing complexity of the organisms . However , in higher eukaryotes there is no correlation between the increased genome size & complexity. This lack of correlation between genome size & genetic complexity is refers to C-value paradox. http://www.ib.bioninja.com.au/_Media/genome-size-graph_med.jpeg Organism Genome size (Mb) S.Cervisiae (yeast) 12 A.Thaliana (mustard plant) 120 D.Melanogaster (fruit fly ) 170 H.Sapiens (humans) 3300 H.Vulgare (barley) 5300 3
- 4. HISTORY OF GENOME SEQUENCING 4
- 5. OUTCOME OF HGP 40+ Models of genomes 2000+ microbial genomes 2000+ viral genomes Projects : HapMap ,1000 genome , encode etc Database & tools for genome analysis : GenBank, HapMap viewer , Genome browser etc Genome tools : NGS, RNAseq , ChipSeq , Genome array etc. https://media.nature.com/m685/nature-assets/nature/journal/v458/n7239/images/nature07943-f3.2.jpg 5
- 6. COST FOR SEQUENCING https://www.genome.gov/27541954/dna-sequencing-costs-data/ Quality Genome coverage 6
- 7. NEXT GENERATION SEQUENCING Strategy : One single reaction for the simultaneous analysis of different genes. Use : Analysis of undifferentiated mutations in heterogeneous diseases. Personalised medicines ,Genetic diseases ,clinical diagnostics . Benefit : Highly cost effective & efficient by simultaneous & fast analysis. Disadvantage : Interdependency of the abundance of data challenging high coverage needed for accuracy. Technologies : 454 , illumina, PacBio , ABI, Helicos, ion Torrent , Nanopores . 7
- 8. 8 Four main DNA sequencing methods 1)Pyrosequencing 2)Sequencing by synthesis 3)Sequencing by ligation 4)Ion semiconductor sequencing https://image.slidesharecdn.com/introductiontonextgenerationsequencingv2-110927082709- phpapp02/95/introduction-to-next-generation-sequencing-16-728.jpg?cb=1317179320 Technologies : 454 , illumina, PacBio , ABI, Helicos, ion Torrent , Nanopores .
- 9. FOUR STEPS IN DATA SEQUENCING 1 ) Library preparation 2) Cluster generation 3) Sequencing 4) Data analysis 9
- 10. Shotgun sequencing by ION Torrent (3 hours) Personal Genome Machine or 454 10 Fragmented DNA adaptors Barcodes Bead oligosides Adapter complement sequences Beads are placed in the wells of micro slides ,amplified by PCR ,dNTPs are flurocent coated & added each at a time. 1 million or 1 billion H Luciferase , pico titer plates P
- 11. Applications Cancer genomics Pre-natal diagnosis 1000 Genomics project Personalised medicines Nanopore 11 https://tse1.mm.bing.net/th?id=OIP.GR5uNIbqUz3lNCYggxl9xAHaDt&pid=15.1&P=0&w=308&h=155
- 12. 12 Thank You Citation :NPTEL Genomics & Proteomics Shomus Biology