MICROARRAY TECHNOLOGY

BySehreen Saddique

Ibtesam Riaz Bakhtawer NasrullahUniversity of Gujrat

OUTLINE

1. Introduction of microarray1. Introduction of microarray

3. Applications of microarray3. Applications of microarray

2. Types of microarray 2. Types of microarray

DNA MICROARRAY

Gene expression

is used to measure the levels of gene expression DNA MICROARRAY

If the hybridised DNA or RNA is labelled fluorescently it can be quantified by scanning of the chip.

DNA microarrays can be manufactured by:

• Photolithography

• Robot spotting

Photolithography

• Each probe 25 bp long• 22-40 probes per gene• Perfect Match (PM) as

well as MisMatch (MM) probes

Robot Spotting

DNA MICROARRAY TECHNIQUE

• Presynthesized DNA strands or synthesis of oligonuleotide sequence

• Spotting of DNA strands to plastic plate or glass plate

DNA MICROARRAY TECHNIQUE

• The two samples to be compared are grown/acquired. In this example treated sample (case) and untreated sample (control).

• Extraction of mRNA material from cells

• Formation of complementary DNA through reverse transcription

• Labeling with fluorescent dyes with Cy3 or Cy5

• Mixing of labeled samples & presenting to microarray spots again by spotters

• Hybridization to a DNA Microarray

• Scanning the Hybridized Array

ANALYSIS OF GENE EXPRESSION DATA

• Analyzing issues

• Different environmental conditions

TYPES OF MICROARRAY

SEHREEN SADDIQUE

ROLL NO:08030614-045

Types of microarray

• There are two types of DNA microarray

• c DNA

• Oligoneuclotide array

c DNA MICROARRAY

1. Prepare sample.

Test Reference

2. Label with fluorescent dyes.

3. Combine cDNAs.

4. Print microarray.

5. Hybridize to microarray.

6. Scan.

Nelzo ErefulCrop Bioinformatics

cDNA Array Sample Preparation

APPLICATIONS OF MICROOARRAY

BAKHTAWER

ROLL:08030614-047

Applications of microarray

Drug Discovery and development

Evolutionary and ecological

genomics

Tiling array Tumor classification

Gene expression

(stress)

Microarray

SNP detection

Applications of microarray

Application in ecology

Microarray and asthma

Disease discovery

Medical application of microarray analysis

treatment of cancer may be tailored to the needs of that individual’s gene expression profile.

Tumor classification

Quackenbush 2006; Retrieved from http://content.nejm.org/cgi/content/full/354/23/2463

tolerant ATGTGTGGAGGAGAAGTGATCCCCGCCGACATGCCGGCGGCGCCGTTCACGCCACGCCAC 60 susceptible ------------------------------ATGCCGGCGGCGCCGTTCACGCCACGCCAC 30 ******************************

tolerant GGCGACGGCGAGACATGGGTTGACAGAAAGAGGAGGAACAAGAAGAAGAGGAAGCGCGGC 120 susceptible GGCGACGGCGAGACATGGGTTGACAGAAAGAGGAGGAACAAGAAGAAGAGGAAGCGCGGC 90 ************************************************************

tolerant GCCGACGAAGAATGGGAGGCCGCCTTCCAGGAGTTCATGGCTGCTGACGACGACGACGAC 180 susceptible GCCGACGAAGAATGGGAGGCCGCCTTCCAGGAGTTCATGGCTGCTGACGACGACGACGAC 150 ************************************************************

tolerant GGCGGCGGACTCGTGTTAAGTAGTAAATCTTTGGTGTTGAGGTCACCAGGTGAAAATGAT 240 susceptible GGCGGCGGACTCGTGTTAAGTAGTAAATCTTTGGTGTTGAGGTCACCAGGTGAAAATGAT 210 ************************************************************

tolerant GCAGGCCGGGGCGCCGCCGCCACCATGTCCATGCCGCTGGACCCCGTGACCGAGGAGGCC 300 susceptible GCAGGCCGGGGCGCCGCCGCCACCATGTCCATGCCGCTGGACCCCGTGACCGAGGAGGCC 270 ************************************************************

tolerant GAGCCGGCGGTGGCTGAGAAGCCTCGCCGGCGCCGGCCGAGGCGGAGCTACGAGTACCAC 360 susceptible GAGCCGGCGGTGGCTGAGAAGCCTCGCCGGCGCCGGCCGAGGCGGAGCTACGAGTACCAC 330 ************************************************************

tolerant GGCATCCGGCAGCGGCCGTGGGGGCGGTGGTCGTCGGAGATCCGCGACCCCGTCAAGGGC 420 susceptible GGCATCCGGCAGCGGCCGTGGGGGCGGTGGTCGTCGGAGATCCGCGACCCCGTCAAGGGC 390 ************************************************************

tolerant GTCCGCCTCTGGCTCGGCACCTTCGACACCGCCGTCGAAGCCGCGCTCGCCTACGACGCC 480 susceptible GTCCGCCTCTGGCTCGGCACCTTCGACACCGCCGTCGAAGCCGCGCTCGCCTACGACGCC 450 ************************************************************

tolerant GAGGCCCGCCGCATCCACGGCTGGAAAGCCCGGACAAACTTCCCACCCGCCGATCTTTCT 540 susceptible GAGGCCCGCCGCATCCACGGCTGGAAAGCCCGGACAAACTTCCCACCCGCCGATCTTTCT 510 ************************************************************

tolerant TCGCCGCCGCCGCCGTCGCAGCCGCTCTGCTTCTTGCTCAACGACAACGGCCTCATCACA 600 susceptible TCGCCGCCGCCGCCGCCGCAGCCGCTCTGCTTCTTGCTCAACGACAACGGCCTCATCACA 570 *************** ********************************************

tolerant ATCGGAGAAGCGCCGACCGACGACGCCGCGTCGACGTCGACGTCGACGACGGAGGCGTCC 660 susceptible ATCGGAGAAGCGCCGACCGACGACGCCGCGTCGACGTCGACGTCGACGACGGAGGCGTCC 630 ************************************************************

tolerant GGCGACGCGCGCATACAACTGGAGTGCTGCTCGGACGACGTGATGGACAGCCTCCTCGCC 720 susceptible GGCGACGCGCGCATACAGCTGGAGTGCTGCTCGGACGACGTGATGGACAGCCTCCTCGCC 690 ***************** ******************************************

tolerant GGCTACGACGTGGCCAGCGGCGACGACATATGGACATGGACATCTGGAGCCTCCTCCACC 780 susceptible GGCTACGACGTGGCCAGCGGCGACGACATATGGACATGGACATCTGGAGCCTCCTCCACC 750 ************************************************************

tolerant TCTGTTAACCAAGAGATCAAGACCCCATCGATCCACCAAAACATATCATATGCAGGTGCC 840 susceptible TCTGTTAACCAAGAGATCAAGACCCCATCGATCCACCAAAACATATCATATGCAGGTGCC 810 ************************************************************

tolerant CGCCCCATGACTTGTCACTTTAAGAATCATAAAAACACTTTTGTACAAATGGAGTGCTCA 900 susceptible CGCCCCATGACTTGTCACTTTAAGAATCATAAAAACACTTTTGTACAAATGGAGTGCTCA 870 ************************************************************

tolerant ACCATGCTAAACTTACTCAAAGGCCACAAACAATAA 936 susceptible ACCATGCTAAACTTACTCAAAGGCCACAAACAATAA 906 ************************************

CCGTCGC

CCGCCGC

tolerant ATGTGTGGAGGAGAAGTGATCCCCGCCGACATGCCGGCGGCGCCGTTCACGCCACGCCAC 60 susceptible ------------------------------ATGCCGGCGGCGCCGTTCACGCCACGCCAC 30 ******************************

tolerant GGCGACGGCGAGACATGGGTTGACAGAAAGAGGAGGAACAAGAAGAAGAGGAAGCGCGGC 120 susceptible GGCGACGGCGAGACATGGGTTGACAGAAAGAGGAGGAACAAGAAGAAGAGGAAGCGCGGC 90 ************************************************************

tolerant GCCGACGAAGAATGGGAGGCCGCCTTCCAGGAGTTCATGGCTGCTGACGACGACGACGAC 180 susceptible GCCGACGAAGAATGGGAGGCCGCCTTCCAGGAGTTCATGGCTGCTGACGACGACGACGAC 150 ************************************************************

tolerant GGCGGCGGACTCGTGTTAAGTAGTAAATCTTTGGTGTTGAGGTCACCAGGTGAAAATGAT 240 susceptible GGCGGCGGACTCGTGTTAAGTAGTAAATCTTTGGTGTTGAGGTCACCAGGTGAAAATGAT 210 ************************************************************

tolerant GCAGGCCGGGGCGCCGCCGCCACCATGTCCATGCCGCTGGACCCCGTGACCGAGGAGGCC 300 susceptible GCAGGCCGGGGCGCCGCCGCCACCATGTCCATGCCGCTGGACCCCGTGACCGAGGAGGCC 270 ************************************************************

tolerant GAGCCGGCGGTGGCTGAGAAGCCTCGCCGGCGCCGGCCGAGGCGGAGCTACGAGTACCAC 360 susceptible GAGCCGGCGGTGGCTGAGAAGCCTCGCCGGCGCCGGCCGAGGCGGAGCTACGAGTACCAC 330 ************************************************************

tolerant GGCATCCGGCAGCGGCCGTGGGGGCGGTGGTCGTCGGAGATCCGCGACCCCGTCAAGGGC 420 susceptible GGCATCCGGCAGCGGCCGTGGGGGCGGTGGTCGTCGGAGATCCGCGACCCCGTCAAGGGC 390 ************************************************************

tolerant GTCCGCCTCTGGCTCGGCACCTTCGACACCGCCGTCGAAGCCGCGCTCGCCTACGACGCC 480 susceptible GTCCGCCTCTGGCTCGGCACCTTCGACACCGCCGTCGAAGCCGCGCTCGCCTACGACGCC 450 ************************************************************

tolerant GAGGCCCGCCGCATCCACGGCTGGAAAGCCCGGACAAACTTCCCACCCGCCGATCTTTCT 540 susceptible GAGGCCCGCCGCATCCACGGCTGGAAAGCCCGGACAAACTTCCCACCCGCCGATCTTTCT 510 ************************************************************

tolerant TCGCCGCCGCCGCCGTCGCAGCCGCTCTGCTTCTTGCTCAACGACAACGGCCTCATCACA 600 susceptible TCGCCGCCGCCGCCGCCGCAGCCGCTCTGCTTCTTGCTCAACGACAACGGCCTCATCACA 570 *************** ********************************************

tolerant ATCGGAGAAGCGCCGACCGACGACGCCGCGTCGACGTCGACGTCGACGACGGAGGCGTCC 660 susceptible ATCGGAGAAGCGCCGACCGACGACGCCGCGTCGACGTCGACGTCGACGACGGAGGCGTCC 630 ************************************************************

tolerant GGCGACGCGCGCATACAACTGGAGTGCTGCTCGGACGACGTGATGGACAGCCTCCTCGCC 720 susceptible GGCGACGCGCGCATACAGCTGGAGTGCTGCTCGGACGACGTGATGGACAGCCTCCTCGCC 690 ***************** ******************************************

tolerant GGCTACGACGTGGCCAGCGGCGACGACATATGGACATGGACATCTGGAGCCTCCTCCACC 780 susceptible GGCTACGACGTGGCCAGCGGCGACGACATATGGACATGGACATCTGGAGCCTCCTCCACC 750 ************************************************************

tolerant TCTGTTAACCAAGAGATCAAGACCCCATCGATCCACCAAAACATATCATATGCAGGTGCC 840 susceptible TCTGTTAACCAAGAGATCAAGACCCCATCGATCCACCAAAACATATCATATGCAGGTGCC 810 ************************************************************

tolerant CGCCCCATGACTTGTCACTTTAAGAATCATAAAAACACTTTTGTACAAATGGAGTGCTCA 900 susceptible CGCCCCATGACTTGTCACTTTAAGAATCATAAAAACACTTTTGTACAAATGGAGTGCTCA 870 ************************************************************

tolerant ACCATGCTAAACTTACTCAAAGGCCACAAACAATAA 936 susceptible ACCATGCTAAACTTACTCAAAGGCCACAAACAATAA 906

************************************

CCGTCGC = tolerant (S186)

CCGCCGC = intolerant (P186)

ACAACTG = tolerant

ACAGCTG = intolerant

SNP

SNP detection using Microarray

Generating DNA Sequence

base callingquality clippingvector clippingcontig assembly

chromatogram files

automatedsequencer

software pipeline

>GENE01

ACCTGTCAGTGTCAACTGCTTCAATAGCTAATGCTAGGCTCGATAATCGCTGGCCTCAGCTCAGTCTAGCATTACGATTACGGAGACCTATGCTTTAGCTAGTAGGAACCTCAGCTCAGTACCTGTCAGTGTCAACTGCTTCAATAGCTAATGCTACTC

>GENE01

ACCTGTCAGTGTCAACTGCTTCAATAGCTAATGCTAGGCTCGATAATCGCTGGCCTCAGCTCAGTCTAGCATTACGATTACGGAGACCTATGCTTTAGCTAGTAGGAACCTCAGCTCAGTACCTGTCAGTGTCAACTGCTTCAATAGCTAATGCTACTC

output