MICROARRAY TECHNOLOGY By Sehreen Saddique Ibtesam Riaz Bakhtawer Nasrullah University of Gujrat

OUTLINE 1. Introduction of microarray 2. Types of microarray 3. Applications of microarray My purpose of showing you the Omics organization is to locate where transcriptomics is and the interrelationships among the various “omics” organizations.

DNA MICROARRAY

Gene expression DNA MICROARRAY is used to measure the levels of gene expression

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

SEHREEN SADDIQUE ROLL NO:08030614-045 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. 4. Print microarray. Test Reference 2. Label with fluorescent dyes. 5. Hybridize to microarray. 3. Combine cDNAs. 6. Scan.

Nelzo Ereful Crop Bioinformatics

cDNA Array Sample Preparation

APPLICATIONS OF MICROOARRAY BAKHTAWER ROLL:08030614-047

Applications of microarray Evolutionary and ecological genomics Drug Discovery and development SNP detection Microarray Gene expression (stress) Tiling array Tumor classification

Applications of microarray

Application in ecology

Disease discovery Microarray and asthma

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

SNP SNP detection using Microarray CCGTCGC = tolerant (S186) CCGTCGC 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 ************************************ 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 ************************************ SNP CCGTCGC = tolerant (S186) CCGCCGC = intolerant (P186) CCGTCGC CCGCCGC ACAACTG = tolerant ACAGCTG = intolerant

Generating DNA Sequence automated sequencer chromatogram files software pipeline base calling quality clipping vector clipping contig assembly output >GENE01 ACCTGTCAGTGTCAACTGCTTCAATAGCTAATGCTAGGCTCGATAATCGCTGGCCTCAGCTCAGTCTAGCATTACGATTACGGAGACCTATGCTTTAGCTAGTAGGAACCTCAGCTCAGTACCTGTCAGTGTCAACTGCTTCAATAGCTAATGCTACTC

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