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Hello to all,
I am new to Q-PCR.
When I put Q-PCR first time for the newly synthesized cDNA, I did not get much difference in cT value between the replicates (of same sample) and also in housekeeping genes cT values of different samples. As I need to analyze the cDNA sample for four different primers, I freeze thawed cDNA for around 10 times and now I am getting difference in cT value (cT1 – 23; cT2 – 28 of same cDNA sample) and also in housekeeping genes cT value among different samples. I tried to reduce pippetting and manual error. Is it due to cDNA regradation? If so, why am I getting difference between cT values of the same sample? (I did duplicate for each sample). Please suggest me.
Thanks,
ram.
