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) Homlogous genes

Orthologs versus paralogs

Microarray Analysis and an example of its application

Genomic DNA Transcriptome Proteome

DATA BASE

Overview

The Human Genome Project

June 2002

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

Map-based Cloning of Human Genome

“Partial Restriction Digests”

Smaller pieces for sequencing

Time Consuming Process whereby: Areas of overlap

between identifies the sequence of DNA

http://www.hhmi.org/biointeractive/dna/DNAi_human_genome_seq.html

Shot-Gun Sequencing of the human genome

http://bcs.whfreeman.com/thelifewire/content/chp17/1702002.html

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

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

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

http://hapmap.ncbi.nlm.nih.gov/

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

Forensic Analysis: STRs FBI: Combined DNA

Index System

13 loci

STRs (4-5 nucleotide repeats)

Example of D1S80 STR Distance migrated on

gel electrophoresis corresponds to size of repeating nucleotide sequence.

cDNA mRNA

Reverse transcriptase

cDNA

Predicting Function from Homology Search

Phylogenetic Profile

Computational method prediction Paralog

Ortholog

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

Microarray Protocol DNA or cDNA is added

robotically to chip

Chips exposed to labeled cDNA

Label monitored by laser

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 haverecurring cancer. Now return to the original microarrays and compare thesetwo groups of women (disease-free and recurring cancer). Is gene expression different?

The composite microarray Can you see any differences?

Recurring cancers

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

Cancer free

Expression patterns varies between patients

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

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

BLAST searches Unknowns and examples of how to search

database provided tomorrow in lab.

Mouse: Mus musculus 2.6 billion bp

99% in common with human genome