Presentation on theme: "Bacterial Physiology (Micr430)"— Presentation transcript:
1 Bacterial Physiology (Micr430) Lecture 13Bacterial Genomics and Functional Genomics(from various sources)
2 Genome and ProteomeGenome: The complete genetic information carried by an organismProteome: Encompasses all the expressed proteins of an organism (protein array)Gene array: an orderly array of sequence tags each specific for one gene of a genome; (other names: gene chip, DNA microchip)
3 Genomics and Functional Genomics Genomics: The study of all of the nucleotide sequences, including structural genes, regulatory sequences, and noncoding DNA segments, in the chromosomes of an organismFunctional genomics: the branch of genomics that determines the biological function of the genes and their products
5 Gene Array or DNA microaray There are two main applicationsIdentification of gene or mutationDetermination of expression level of genesThere are two formats:cDNA (500 – 5000 bp) molecules are spotted on the chipArray of oligonucleotides are synthesized and immobilized on the chip
6 Gene Array or DNA microarray 1) Probe (cDNA/oligo with known identity)2) Chip fabrication (Putting probes on the chip)3) Target (fluorecently labeled sample)4) Assay5) Readout6) Informatics
9 DNA microarray applications Gene discoveryStudy of gene functions and expressionStudy relationship of genesDisease diagnostics – infectious diseases
10 Study of gene functionGene replacement (knock-out): also known as reverse genetics. The purpose is to remove (knock-out) most of one gene and see what happens to the phenotype of the organism. Suicide vector is usedTransposon mutagenesis. Similar to above but is more random disruption of genes.
12 Study of gene functionAntisense RNA expression. Random cloning and expression of short pieces of genomic DNA on a plasmid in an microorganism to elucidate the function of the genesEssential gene: a gene whose function is absolutely required for the growth and survival of a cell.
13 Conditional Antisense Inhibition of Protein Synthesis Inducible promoterNormal cellAntisense cellPlasmid DNANoproteinProteinAntisense RNAXmRNAmRNADNADNA
14 Shotgun Antisense Expression Determines Essentiality of Genes Pathogen genomeMillions of random DNA fragmentsNon essential geneblocked by antisenseEssential geneNo cell growthmRNAEssential ProteinDNA
15 Ultra-Rapid Functional Genomics Identify >100 essential gene drug targets per monthAntisense(+ inducer)No antisense(- inducer)
16 Traditional ToolsReporter gene: A gene whose product can be easily assayedTranscriptional gene fusionsTranslational gene fusionsPCRDNA Mobility Shift