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Genomic Analysis
Chapter 19
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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
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cDNA can also be studiedwww.bio.davidson.edu/Courses/genomics/RTPCR/RT_PCR.html :
cDNA:
mRNA Reverse transcriptase
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SEQUENCING DNA
Sanger-Dideoxymethod
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Sanger Method dideoxynucleotides: lack a 3’ OH
But have the 2’ and 3’ have hydrogen
NO DNA EXTENSION
Dideoxy key point!
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DNA PCR amplification is performed prior to sequence analysis
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Autoradiogram
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Can you sequence without using radioactivity?
Radioactive!
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Yes: Example of Labelled Probes ddGTP ddATP ddTTP ddCTP
Orange Black Red Blue
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Microarray Analysis
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Microarray Protocol DNA or cDNA is added
robotically to chip
Chips exposed to labeled cDNA
Label monitored by laser
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Example of: Microarrays from breast tumors cDNA from breast tumors from 2 groups of
patients
Metastatic Cancer Disease Free
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Microarrays of 70 genes
Metastatic cancers
(row is a patient’s tumor, a column a gene)
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Expression patterns varies between patients
Those “genes” that are green from “normal” tissue genes
Those “genes” that are red are associated with cancer!
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Genomic DNA Transcriptome Proteome
DATA BASE
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Human Genome ProjectJune, 2002
Dr. Craig Venter
President Clinton
Dr. Francis Collins
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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
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Mouse: Mus musculus 2.6 billion bp
99% in common with human genome
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Genomic Projects: provide us information about:
Expressed-sequence tags (ESTs)
Single-nucleotide polymorphisms (SNPs) & Haplotypes
Variable Number Tandem Repeats (VNTRs)
Need to understand cDNA
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Haplotype:A set of SNPs and other variants on the same chromosome
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Variable number tandem repeats(VNTRs) in multiple loci
Short stretches of DNA base pairs that are repeated at multiple loci in the chromosomes.
GAGGAGGAG
GAGGAGGAGGAGGAGGAG
GAGGAGGAGGAGGAGGAGGAGGAGGAGGAGGAGGAGGAG
GAGGAGGAGGAGGAGGAG
GAGGAGGAGGAGGAG
GAGGAGGAGGAG
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Predicting Function from Homology Search (only topic with additional slide )
Phylogenetic Profile
Gene Neighbor Analysis
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Computational method prediction Paralog
Ortholog
Paralogs: Homologous sequences in same species Orthologs
Homologous Sequences in Different species
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