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M.SHANMUGA PRIYAA
M.Sc BIOTECHNOLOGY
GENOME INSTITUTE OF SINGAPORE
Comparative study, on the expression of protein
kinase genes pertaining to the signaling pathway of
the cell division cycle (cdc), of S.pombe in both the
wild and mutant strains is being investigated using
micro array technology.
• Schizosaccharomyces pombe-
fission yeast.
• S.pombe is,
an unicellular eukaryote,
rod-shaped
2 - 3 µm in diameter
7-14 µm in length
WILD MUTANT
CELL CYCLE :
INTERPHASE
G1-S-G2
M PHASE
PROPHASE,
METAPHASE,
ANAPHASE,
TELOPHASE.
Key players in the progression of the cell cycle are a group of kinases.
KINASES
• Binds to cyclins-regulate
cyclins kinase activity.
• Inactive kinases-cdk’s
• Add phosphate group
• Phosphorylation of proteins at different stages of the cell cycle is
necessary for the cycle to complete.
GENE EXPRESSION :
Complex pattern of expression
Determines the characteristics of a cell and its role in the organism.
Abnormal pattern- cancer.
MICROARRAY:
• To assay gene expression within a single cell.
• To compare gene expression in two different cell types or tissue samples.
• To examine the expression of hundreds or thousands of genes at once.
STEPS:
1.Preparation of DNA chips using known DNAs.
2.Generation of a hybridization solution containing a mixture of
fluorescently labeled cDNAs (mutant type).
3.Incubation of the hybridization mixture containing fluorescently labeled cDNAs with
DNA chip.
4.Detection of bound cDNAs using laser technology and storing data in a computer.
5.Analyzing and interpreting the data using computational methods.
Cell culture
Spotting of array’s
Hybridization
Scanning
•The image analysis - 2 final intensity measurements for the red and green
are obtained for every single spot.
SCANNED SLIDE
• APPLICATION
- tissue/cell phenotype analysis by multiple gene expression assay,
- identification of pathogens,
- genome-wide mutation survey
- diagnosis of genotypic polymorphisms and mutations.
- drug discovery,
- developmental biology
- molecular analysis of various diseases.