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 GenomicsSequencing Genome (Reference)

Functional GenomicsGene Expression

Proteomics

MetabolomicsComparative Genomics

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Genomic libraries

DNA Libraries

cDNA libraries[for EST sequencing][for Genome sequencing]

ESTs sequences

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 gene5

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

• Ref = Koonin, E.V. and Galperin, M.Y. 2004 In: Sequence-Evolution-Function. p.196

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Requiring Bioinformatics

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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