QPCR versus regular PCR for Chip - (Nov/01/2007 )

Hi,

My project is to examine whether a transcription factor binds with the promoter region of a target gene. So, it's basically a "yes" or "no" question. While I read the papers, I saw a lot of people are using regular PCR (agarose gel and EB detection) for Chip detection . But the other lab in our deparment is using QPCR for all of their Chip experiments and it looks like that everyone in this forum is also using QPCR.
So, I am wondering whether I can use regular PCR instead of QPCR for my purpose . Regular PCR are more convenient and cheaper than real time PCR in our lab, because our lab don't have QPCR machine and have to pay a lot money to let a genomic facility to run the QPCR for us.

Thanks

Semi-quantitative PCR can be used for ChIP.
The only drawback is that it is not quantitative and
if you do not get an "on/off" answer it is more confusing than QPCR.
The impt. parameters for semiquantitative are as follows:
#1 Make sure your primers amplify in linear range. Do not Saturate pull down products!
#2 ALWAYS USE NEGATIVE CONTROLS.
#3 ALWAYS USE POSITIVE CONTROLS.
#4 Did I mention positive and negative controls?


When doing semiquantitative the results are usually clear when your bands
are compared directly (on the same gel) to positive and negative controls.

Hi,

for ChIP I do both quantitative and "normal" PCR. Normal PCR is to check whether I have signal or not and whether my inputs are in equal amount. If I am comparing the amounts of DNA (i.e. in a time course experiment), then I do real-time PCR as well. There I can normalize my samples to the inputs and express amount of samples as fold-change. In normal PCR, it is also possible that I have a lot DNA as starting material and the signal reaches a plateau (then it wouldn`t be possible to compare my samples). There I use real-time PCR as well.

Cheers