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DNA Microarray Technology
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DNA Microarray TechnologyUnder the guidance of
Dr .Manjunath
Ankitha Hirematha3rd sem, MSc.,
Dept. of BiotechnologyKuvempu university.
CONTENTS
• Introduction• Historical Background• Principle• Overview steps• Preparation of slide• Microarray scanning• Data analysis and normalization
DNA Microarray Technology
Why ?
To analyze the expression of thousands of genes in single reaction, very quickly and in an efficient manner.
To understand the genetic causes for the abnormal functioning of the human body.
To understand which genes are active and which genes are inactive in different cell types.
What is DNA Microarray Technology ?
It is “an orderly arrangement of thousands of identified sequenced genes printed on an impermeable solid support , usually glass, silicon chips or nylon membrane”.
DNA microarray chips are also known as DNA chips, DNA arrays, or biochips.
Historical background :
• Southern blotting was developed in the year 1975.
Sir Edwin Southern
• The concept of DNA microarrays began in the mid 1980s.
• Pin based robotic system was developed by Lehrach’s group in 1990.
•“Quantitative Monitoring of Gene Expression Patterns with a complementary DNA microarray” reported by Patrick Brown, Mark Schena and colleagues in Science (1995).
• Steve Fodor developed scanner for reading the output.
Sir Steve Fodor
Sir Patrick Brown
•Mark Schena was proclaimed as the “Father of Microarray Technology”.
Mark Schena
Principle of DNA Microarray:
An overview of steps involved in DNA Microarray
Preparation of DNA Microarray Slide :
The solid supports used are glass, silicon or nylon membranes
Length of slide is 25 X75mm
Within the area of 3.6cm2 , 10,000 to 20,000 spots(genes)
Diameter of spot is 50-150μm
Distance between the spots is 200-250 μm.
Fabrication
• Photolithography• Robot spotting• Inkjet
Photolithography
Robot Spotting
Inkjet spotting
Performing DNA Microarray Experiment
Eg. Saccharomyces cerevisiae
O2 O2
Centrifuge
Supernatant is discarded
Add extraction buffer
Remove that buffer containing mRNA and place in fresh tube
AAAAAAAAAAA
AAAAAAAAAAA
AAAAAAAAAAA
AAAAAAAAAAA
AAAAAAAAAAA
AAAAAAAAAAA
Reverse Transcription
TTTTTTTTTTTTT
TTTTTTTTTTTTT
TTTTTTTTTTTTTTTTTTTTTTTTTT
TTTTTTTTTTTTT
TTTTTTTTTTTTT
c DNA
Mix
GATC
GATC
GATC
ATGC
ATGC
ATGC
TCAG
TCAG
TCAG
SCANNED
When green laser comes,
When red laser comes,
Merged image :
DNA Microarray Scanner
• Fluorescent intensity is measured
Data analysis and normalization
“Normalization means to adjust the microarray data for effects which arise from variation in the technology”.
Applications
• Gene expresion profiling, SNP detection, human diseases etc..
• IPSC (Induced Pleuripotent Stem Cell) cell lines are validated and monitored.
• Toxic studies
CONCLUSION
Since this DNA microarray technology is used for the analysis of expression of thousands of genes at once, and has wide applications in in analyzing various diseases. It is one of the smartest technique.
•Tim Lenoir ,Eric Giannella (2006) The emergence and diffusion of DNA microarray technology. Journal of Biomedical Discovery and Collaboration(JBDC). 1:11
•Samuel K. Moore (2001) Biochips are now a critical tool for analyzing the human genome--and a lucrative product attracting technology giants. IEEE spectrum• http://www.premierbiosoft.com/tech_notes/microarray.html
•http://www.genome.gov/10000533
•http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1590052/
•http://www.medscape.com/viewarticle/543871_2
•http://en.wikipedia.org/wiki/DNA_microarray#History
•http://grf.lshtm.ac.uk/microarrayoverview.htm#m1
•http://www.ncbi.nlm.nih.gov/About/primer/microarrays.html
•http://www.digizyme.com/portfolio/microarraysfab/photolith.html
REFERENCES
Thank You