2. Genomic Analysis Chapter 19

3. Overview of topics to be discussed  How to sequence genomic DNA (we will have to touch briefly on polymerase chain reaction—a topic from our upcoming “PCR” lab on 5/15). cDNA  Microarray Analysis  The Human Genome Project  How to use information from any genome sequence: Single Nucleotide Polymorphisms Predicting Function from Sequence

4. cDNA can also be studiedwww.bio.davidson.edu/Courses/genomics/RTPCR/RT_PCR.html :  cDNA: mRNA Reverse transcriptase

5. SEQUENCING DNA Sanger-Dideoxymethod

7.  Sanger Method dideoxynucleotides: lack a 3’ OH  But have the 2’ and 3’ have hydrogen  NO DNA EXTENSION Dideoxy key point!

8. DNA PCR amplification is performed prior to sequence analysis

11. Autoradiogram

12. Can you sequence without using radioactivity? Radioactive!

13. Yes: Example of Labelled Probes  ddGTP  ddATP  ddTTP  ddCTP  Orange  Black  Red  Blue

15. Microarray Analysis

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

19. Example of: Microarrays from breast tumors  cDNA from breast tumors from 2 groups of patients Metastatic CancerDisease Free

20. Microarrays of 70 genes Metastatic cancers (row is a patient’s tumor, a column a gene)

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

23. Genomic DNATranscriptome Proteome DATA BASE

24. Human Genome Project June, 2002 Dr. Craig Venter President Clinton Dr. Francis Collins

25. The initial draft of the Human Genome 3.4 billion base pairs 32,000 genes Average gene 27,000 bases Chromosome 17, 19, 22 high density May encode 96,000 proteins

26. Mouse: Mus musculus  2.6 billion bp  99% in common with human genome

27. Genomic Projects: provide us information about:  Expressed-sequence tags (ESTs)  Single-nucleotide polymorphisms (SNPs) & Haplotypes  Variable Number Tandem Repeats (VNTRs) Need to understand cDNA

28. Haplotype:A set of SNPs and other variants on the same chromosome

29. 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 GAGGAGGAGGAG GAGGAGGAG GAGGAGGAGGAGGAGGAG GAGGAGGAGGAGGAG GAGGAGGAGGAG

30. Predicting Function from  Homology Search (only topic with additional slide )  Phylogenetic Profile  Gene Neighbor Analysis

31. Computational method prediction  Paralog  Ortholog Paralogs: Homologous sequences in same species Orthologs Homologous Sequences in Different species