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Genomics Analysis Chapter 20

Genomics Analysis Chapter 20. Overview of topics to be discussed. The Human Genome Analysis Variable Number Tandem Repeats Short Tandem Repeats Single Nucleotide Polymorphisms and Haplotypes Expressed sequence tags (ESTs) Predicting Protein Function BLAST searches (in lab)

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Genomics Analysis Chapter 20

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  1. Genomics Analysis Chapter 20

  2. Overview of topics to be discussed • The Human Genome Analysis • Variable Number Tandem Repeats • Short Tandem Repeats • Single Nucleotide Polymorphisms and Haplotypes • Expressed sequence tags (ESTs) • Predicting Protein Function • BLAST searches (in lab) • Homlogous genes • Orthologs versus paralogs • Microarray Analysis and an example of its application

  3. Overview Genomic DNA Transcriptome Proteome DATA BASE

  4. The Human Genome Project June 2002

  5. Methods to used include: • The two strategies used: • Bac-to-Bac Sequencing (Map Based Sequencing) • Shot-Gun Method http://www.sciencemag.org/content/291/5507/1304.full

  6. Map-based Cloning of Human Genome “Partial Restriction Digests” Smaller pieces for sequencing

  7. Time Consuming Process whereby: • Areas of overlap between identifies the sequence of DNA http://www.hhmi.org/biointeractive/dna/DNAi_human_genome_seq.html

  8. Shot-Gun Sequencing of the human genome • http://bcs.whfreeman.com/thelifewire/content/chp17/1702002.html

  9. Human Genome approximated: 3.4 billion base pairs 32,000 genes Average gene 27,000 bases Chromosome 17, 19, 22 high density May encode 96,000 proteins

  10. Ongoing data bases from human genome project: • Variable Number Tandem Repeats (VNTRs) • Short Tandem Repeats (STRs) • Single-nucleotide polymorphisms (SNPs) SNP Maps and haplotypes • http://www.ncbi.nlm.nih.gov/pubmed/11029002 • http://www.fruitfly.org/SNP/ • Expressed-sequence tags (ESTs) Oops! Need to understand cDNA

  11. Haplotype:A set of SNPs and other varians on the same chromosome http://hapmap.ncbi.nlm.nih.gov/

  12. Variable number tandem repeats(VNTRs) in multiple loci Short stretches of DNA base pairs that are repeated at multiple loci in the chromosomes. GAG GAGGAG GAGGAGGAG GAGGAGGAG GAGGAGGAGGAG GAGGAGGAG GAGGAGGAGGAGGAGGAG GAGGAGGAGGAGGAG GAGGAGGAGGAG GAGGAGGAGGAGGAGGAG

  13. Forensic Analysis: STRs • FBI: Combined DNA Index System • 13 loci • STRs (4-5 nucleotide repeats)

  14. Example of D1S80 STR • Distance migrated on gel electrophoresis corresponds to size of repeating nucleotide sequence.

  15. cDNA • mRNA • Reverse transcriptase • cDNA

  16. Predicting Function from • Homology Search • Phylogenetic Profile

  17. Computational method prediction • Paralog • Ortholog

  18. Microarray Analysishttp://www.youtube.com/watch?v=VNsThMNjKhM

  19. Microarray Protocol • DNA or cDNA is added robotically to chip • Chips exposed to labeled cDNA • Label monitored by laser

  20. Microarrays from breast tumors • cDNA from breast tumors breast cancer patients EACH patient has a chip and microarray containing “candidate” genes Time passes and some women remain “cancer-free” while other women have recurring cancer. Now return to the original microarrays and compare these two groups of women (disease-free and recurring cancer). Is gene expression different?

  21. The composite microarrayCan you see any differences? (Each row is a patient’s tumor, a column a gene) Cancer free Recurring cancers

  22. Expression patterns varies between patients • Those “genes” that are green from “normal” tissue genes • Those “genes” that are red are associated with cancer!

  23. BLAST searches • Unknowns and examples of how to search database provided tomorrow in lab.

  24. Mouse: Mus musculus • 2.6 billion bp • 99% in common with human genome

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