Plant Genomics II - Lecture Notes - Bioinformatics and Genomics | BSCI 411, Study notes of Biology

Download Plant Genomics II - Lecture Notes - Bioinformatics and Genomics | BSCI 411 and more Study notes Biology in PDF only on Docsity! Lecture 9-10. Plant genomics II Functional genomics: Identify the function of each and every gene in the genome. Since the characterization of the function of a protein domain in one organism generally provides hint to its function in another organism, the first goal of functional genomics is to identify as many genes as possible in major model organisms Basic Approaches A. Forward genetics: Random mutagenesis, screen for traits of interests Chromosome walking or transposon-tagging B. Reverse genetics: disrupt a particular gene or set of genes with known seq. C. Fine structure genetics D. Gene expression profile (analyses of transcriptome) B. High-throughput reverse genetics 1. PCR-based screen for T-DNA or transposon insertion mutations in specific genes---Wisconsin knockout facility 2. Database searches--salk institute lines 3. TILLING 4. RNAi (RNA interference) 5. Gain-of-function (activation-tagging) mutagenesis 3. TILLING (Targeting Induced Local Lesions IN Genomes) Arabidopsis Tilling website:http://tilling.fhcrc.org:9366/ EMS Isolated DNA pool Gene-specific primersPCRheat & coolCEL I Denature 1 2 3 4 5 6 7 8 9 10 4. RNA interference (RNAi) loss of gene activity Gene X NPTII 35S enhancer 5. Gain-of-function (Activation tagging) mutagenesis Over-express Gene X Leading to gain-of-function phenotype Gene X GUS Minimal promoter Enhancer of gene X T-DNA 2. Enhancer-trap Dr. Tom Jack’s website http://www.dartmouth.edu/~tjack/ GAL4 Minimal promoter Tissue-specific Enhancer 3. GAL4-mediated over expression X 5X UAS Gene Z www.plantsci.cam.ac.uk/Haseloff/gene_expression/geneExpFrameset.html 5X UAS GFP and/or 4.Yeast Two Hybrid (Y2H) Assay to test interaction between two proteins GAL4 DB GAL4 AD X Y HIS3 or lacZ 5XUAS I. cDNA microarrays Prepare cuNA Rope Pravers Mlleresirrely  II. Oligonucleotide microarrays (Affymetrix GeneChip) o o ooo mask Light deprotection OH OH ooo T oooT T