STANDARDIZATION OF QUANTITATIVE REAL TIME PCR (QRTPCR) FOR DETECTION OF INFLAMMASOMES FOLLOWING EXPOSURE OF PPR VACCINE IN GOATSD.K. SHARMA, B. JOSEPH1, R. SINGATHIA, R. K. NAGDA, M. SHARMA AND S. K. BAROLIA
In present study we standardized the qRTPCR for detection of relative change in gene expression of inflammasome components using gene specific primers. These primers were designed for different components of inflammasome, i.e. NLRP3, ASC, and Caspases1. RT PCR was standardized with cDNA prepared from RNA extracted from PBMCs of goat blood; PBMCs were collected just before PPR vaccination and at different time points 0, 3, 7, 15, 30, 60 and at 90 days post vaccination. We found amplification at specific melting temperature. The amplification of various gene was further confirmed by agarose gel electrophoresis for presence of band of specific size, i.e. 409 bp for Î² actin, 332 bp for NLRP3, 306 bp for ASC, and 209 bp for Caspase-1.
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