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