Multiple reactions to run, each with their own preferred annealing temperature? Grabbed two primers of different Tm’s to amplify a region and trying to work out the best program?
OK, confession time. I used to work some crazy hours in order to fit some library screening into a part time job while finishing my degree. No, that’s not the confession – more, when I put the PCR on I
Designing primers for your high resolution melting is easy enough. A quick run with some known genotypes …. and look for the melting curve differences among samples. Too simple. But what about the temperature-neutral SNP’s? The what?
You’ve done the hard work – you’ve got good quality RNA. You’ve seen ribosomal bands on a gel (or *cough* Fragment Analyzer). A good RQN/RIN (if that’s what you go by) and everything’s great.
Reagent contamination in PCR has long been documented – not only by operators but also from commercial sources long before those operators ever laid DNA-ridden hands on them.
While contamination of some/many Taq polymerase enzymes with residual host E. coli
