Genomics – analysis of entire genomes

Websites:

http://www.bcgsc.bc.ca (BC genome sequence centre)

http://genomics.phrma.org/ (Reference site)

bioinformatics

– computer analysis of genomic information

applications:

medical – human and pathogen genomes

– diagnostics and pharmaceuticals

agriculture – cow, corn, rice genomes

biodiversity & molecular evolution – various genomes

levels of analysis:

genome

– complete genetic material (usually DNA) of organism

studied by: sequencing

transcriptome – complete set of RNA transcripts in cell

– under specific conditions

studied by: “functional genomics”

– expressed parts of genome under specific conditions

common technique: expression profiling (DNA arrays)

– attach different cDNAs (or gene-specific oligos)

– in defined pattern on substrate

– probe with labelled cDNAs

– from reverse transcription of total cell mRNA

(from specific cell type/condition)

– labelled cDNAs – hybridize to matching sequence spots

– amount of hybridization

– shows amount of mRNA present in cell

– same principle as Southern, but clearer results

– substrates may be:

nylon membranes – cheap “homemade” arrays

– good results, but less genes screened at once

glass slides/silica “gene chips”

– commercial manufacturers – expensive

– used in pharmaceutical research

proteome – complete set of proteins in cell

– under specific conditions

studied by: 2-dimensional gel electrophoresis

– traditional technique

– first run isoelectric focusing gel

– separate proteins by charge

– then run through SDS-PAGE at right angle to 1^st

– separate proteins by size

– results show number and amounts of protein in cell

– but not protein interactions (denatured protein)

newer technique: two-hybrid analysis

(guilt-by-association)

– screen for protein interactions

– use sets of fusion protein vectors

(cloned genes expressed as fusions with vector gene)

– fusions with different parts of transcription activator

– 1 set fuses proteins to DNA-binding domain

– other set fuses proteins to activator domain

– if 2 fusions combined in same cell

– functional transcription activator complex may form

– only if products of cloned gene interact

– functional transcription activator

– turns on expression of marker gene phenotype

metabolome – complete set of metabolites in cell

(low molecular weight enzyme substrates/products)

– under specific conditions

study using: mass spectometry, nuclear magnetic resonance spectrophotometry (NMR)