Transcript Powerpoint - University of British Columbia

Genomics
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Genomics relies on high-throughput
technologies
• Automated sequencers
• Robotics
– Colony pickers and Arrayers for sequencing
– Microarray spotters
– Protein Spot-Picker and In-gel proteolytic
digestion
• High-throughput genetics
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Areas of Genomics
Sequencing Genome (Reference)
Gene Expression
Functional Genomics
Proteomics
Comparative
Genomics
Metabolomics
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DNA Libraries
Genomic libraries
[for Genome sequencing]
ESTs
sequences
cDNA libraries
[for EST sequencing]
Full Length cDNA
sequences
• cDNA libraries are needed for EST sequencing
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EST Library
• Expressed Sequence Tags
• Specific tissues and developmental stages = mRNA
isolation (Conversion to cDNA)
• Require cloning of cDNAs
• Require many different tissues = good coverage of
genomic information
• Usually sequence from 5’ or 3’ end (known as pair end or
mate end sequencing)
• Will require more $$ to sequence both ends
• Usually less than 60% of genes coverage
• Will not have regulatory elements information
• Paralogs issue when figuring out the annotation and
function of the gene
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FL cDNA Library
• Full Length cDNA library
• Require intact length of mRNA
• Sequencing from both ends of the cDNA clone
using the vector sequences
• Clone size selection (>800 bases)
• Require finishing of the larger clones by
designing more primers internal to clone insert
• Should translate to a full protein
• Require more $$$ and effort to have the data
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Next Generation(NG)
Sequencing
• Sanger sequencing for the past 25 years
• Expensive, little coverage, limited
nucleotide (nt) per run
• Ngseq (NG 2nd) has greatly reduced cost
per amount of data generated, greater
coverage
• Different technologies pending on
company producing the technology (Metzker
2010 Nature Reviews/Genetics)
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Next Generation(NG)
Sequencing
• NGseq (NG 3rd) has greatly reduced cost
per amount of data generated, greater
coverage
• Much longer reads vs NG 2nd generation
• Different technologies pending on
company producing the technology (Metzker
2010 Nature Reviews/Genetics)
• Ion Torrent, PacBio, Oxford Nanopore
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PacBio NG 3rd
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Genome Annotation
Requiring Bioinformatics
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Ref = Koonin, E.V. and Galperin, M.Y. 2004 In:
Sequence-Evolution-Function. p.196
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Bioinformatics
• Biology, computer science, and information technology
merge = a single discipline
• Creating and maintaining databases to store biological
information eg. nucleotide, amino acid sequences
• Development of complex interfaces whereby researchers
could both access existing data as well as submit new or
revised data
• Develop a comprehensive picture of normal cellular
activities
• Involves analysis and interpretation of various types of
data, including nucleotide and amino acid sequences,
protein domains, and protein structures
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