Genomics and the use of Next Genera3on Sequencing (NGS)
TechniquesIMBB2016
BecA-ILRIHub,NairobiMay9–20,2016
FrancescaStomeo
Genomics
The study of the structure, organization, function, evolution of genomes (including interactions with the environment)
Genomics before NGS
1900 1977
1980 1990
1994-98
1998
2000
Watson and Crick identify DNA (the double helix) as the Chemical basis of heredity
Human Genome Projects (HPG) begins-an international effort to map and sequence all the genes in the human genome
Sanger and Gilbert derive methods of sequencing DNA
Working Draft of the human genome sequencing complete
Human genome project completed 2003
Rediscovery of Mendel's laws helps establish the science of genetics
Methodsofsequencing
• Sangerdideoxymethod(primerextension–termina@on)• Maxam-Gilbertchemicalcleavedmethod(DNAlabelled,chemicallycleavedinsequence-dependentmanner.• NextGenera@onSequencing(pyrosequencing,reversibledye-terminators,etc.)
• SinglestrandedDNAtemplate• AprimerforDNAsynthesis• DNApolymerase• Deoxynucleosidetriphosphates(normaldNTP)–extendsDNAstrand• Dideoxynucleo@detriphosphates(ddNTP)–terminatessynthesis
Cost for sequencing genomes rapidly reduced
What is NGS?
NGS plaBorms
Illumina454
ABISOLiD
General steps of illumina NGS technologies
GenomicDNA/cDNA
GenomicDNA/cDNA
General steps of illumina NGS technologies
GenomicDNA/cDNA
Library
General steps of illumina NGS technologies
General steps of Illumina NGS technologies
Amplifica@onstep
GenomicDNA/cDNA
Library
IlluminaHiSeq2000
454GSFLX SOLiDv4
Readlength 700bp 2*100bp 50*2bpAccuracy 99.9% 98% 99.94%Outputdata 0.7GB 600GB 120GBCost/millionbases $10 $0.07 $0.13
Illumina technologies
Nextera DNANexteraXT smallgenomesTruSeq totalRNATruSeqStranded RNAribosomalRNAremoved
Scope of Sequencing
Metagenomics (viral/bacterial)
Compilation of complete genomes, metagenomes,
annotation and curation of metadata
(meta)genome sequencing
Extraction of important biological
information
Vaccine dvlpmt
Diagnostics
Global diseases surveillance
Drug dvlpmt
Improved Public health intervention
geographical mapping
sequence variation analysis
Primer, microarray
phylogenetic analysis
protein modeling Databases
Improved drug selection
discovery of new micro-
organisms and pathways
Environmental sustainability
From Sequence to impact
Diversity studies
A reference genome for common bean and genome-wide analysis of dual domestications Schmutz et. al 2014 Nature Genetics 46
Expression studies
ThedevelopmentaltranscriptomeofDrosophilamelanogaster
Sequencing ancient genomes
Miller,W.etal.Sequencingthenucleargenomeoftheex@nctwoollymammoth.Nature456,387–390(2008).
Biological Ques3on?
Thank youIMBB2016
BecA-ILRIHub,NairobiMay9–20,2016
FrancescaStomeo