no RT controls - can I run on separate plate and for just one primer set? (Jul/19/2006 )
Hi there everyone
I have run PCRs to investigate the suitability of 6 reference genes for my experimental setup. I have however neglected to run no RT controls on the plates (although i have run no template controls). Would it be acceptable for me to run a separate plate with no RT controls to show the absence of genomic contamination in my samples (yes I know I should really have done this first but all my RNA was DNase treated and I was under the misconception that my primers did not amplify gDNA). Could I do this using just one primer set if I include a positive control?
Any advice is much appreciated.
I think this is okay as long as:
1. your no RT controls are from the same samples you are running with RT (obviously)
2. you do include a positive control, just to assure nothing is wrong with the reaction.
3. you should be able to do one primer set if you are confident it would amplify any contaminating DNA that is there (as I don't know your primers, I am assuming they would), because the real question you are answering here is whether or not you have contaminating gDNA. So, if one of your primer sets tells you this, then that should suffice.