can I use routine PCR to assess whether a gene is expressed or not? - (Apr/26/2010 )
I want to know whether a gene is expressed or not in a bacterium, just expressed or not, and the expression level is not taken into my consideration.
I designed a pair of specific primers for this gene using a RT-PCR primer design software, Beacon Designer. So can I use cDNA as template and the pair of specific primers to run a routine PCR, and if there is band with correct length after electrophoresis, I can conclude that this gene is expressed?
Thanks a lot.
It depends on your definition of "expressed" (we had this conversation somewhere else on the forums once) -- does "expressed" mean "transcribed", or does "expressed" require translation and the production of an active protein?
I think the band just confirms that the corresponding gene is transcribed, right? Thanks.
HomeBrew on Apr 26 2010, 07:10 PM said:
Transcription is a fuzzy process, not ON/OFF like a switch. Therefore, if you have enough cells, you'll get a band for everything, inluding non-transcribed genes. I'd recommend to use a positive and a negative control. Then you can say: compared to gene A, which is usually not transcribed, and similar to gene B, which is usually transcribed... So you'll end up with some kind of transcription level.