Genomic Analysis

Chapter 19

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

cDNA can also be studiedwww.bio.davidson.edu/Courses/genomics/RTPCR/RT_PCR.html :

cDNA:

mRNA Reverse transcriptase

SEQUENCING DNA

Sanger-Dideoxymethod

Sanger Method dideoxynucleotides: lack a 3’ OH

But have the 2’ and 3’ have hydrogen

NO DNA EXTENSION

Dideoxy key point!

DNA PCR amplification is performed prior to sequence analysis

Autoradiogram

Can you sequence without using radioactivity?

Radioactive!

Yes: Example of Labelled Probes ddGTP ddATP ddTTP ddCTP

Orange Black Red Blue

Microarray Analysis

Microarray Protocol DNA or cDNA is added

robotically to chip

Chips exposed to labeled cDNA

Label monitored by laser

Example of: Microarrays from breast tumors cDNA from breast tumors from 2 groups of

patients

Metastatic Cancer Disease Free

Microarrays of 70 genes

Metastatic cancers

(row is a patient’s tumor, a column a gene)

Expression patterns varies between patients

Those “genes” that are green from “normal” tissue genes

Those “genes” that are red are associated with cancer!

Genomic DNA Transcriptome Proteome

DATA BASE

Human Genome ProjectJune, 2002

Dr. Craig Venter

President Clinton

Dr. Francis Collins

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

Mouse: Mus musculus 2.6 billion bp

99% in common with human genome

Genomic Projects: provide us information about:

Expressed-sequence tags (ESTs)

Single-nucleotide polymorphisms (SNPs) & Haplotypes

Variable Number Tandem Repeats (VNTRs)

Need to understand cDNA

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

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

Predicting Function from Homology Search (only topic with additional slide )

Phylogenetic Profile

Gene Neighbor Analysis

Computational method prediction Paralog

Ortholog

Paralogs: Homologous sequences in same species Orthologs

Homologous Sequences in Different species