HEPG2 cells (human hepatocellular liver carcinoma cell line) were incubated for 72 hours with different conditions:
1. Medium containing cholesterol (= control group)
2. Medium without cholesterol
3. Medium containing mevastatin
The effect of these conditions (group 2 + 3) on the expression profile of the LDL receptor will be determined. Therefore, RNA will be isolated from the HEPG2 cells, followed by the synthesis of cDNA. This will be used to measure the expression of LDL receptor (mRNA) using real-time PCR.
I wouldn't use the same well for both...This would be akin to a multiplex PCR basically, I would look into that literature to find potential pitfalls (cons).
qPCR is finicky enough without adding the potential negative interactions of 2 different primer sets having to work in the same well. But I suppose the possibility of it working well is there.
The only Pro I could see is that you would be doing the reaction on EXACTLY the same sample, which I suppose is a bonus. Plus you would save sample by using half as much and some time by pipetting into half as many wells. Otherwise, if you make a mastermix and aliquot into different wells with your respective primers, you are essentially analyzing the same sample anyway, which is what people usually do. Hope this is somewhat helpful.
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