Splice varient quantification in RT real time PCR? - (May/24/2011 )
Am planning to analyse differential expression of mRNA of my protein of interest by RT real time pcr in treated and untreated cell line.
Problem arised when i checked primers in convential PCR, i found 4 pcr products of different size, sequencing revelaed they are splice varient.
my question: is it advicable to use the same primer which gave 4 pcr product in real time pcr (sybr green based). if not what will be alternative method to quantify my gene of interest.
You need to decide if you want to quantify all splice variants or only some of them. Each creates different protein. If you want to quantify only one splice variant you have to design new primers.