Real time (CT IN NEGETIVE WELL AND DOBLE MELTING CURVE) - (Sep/02/2013 )
i have working on rbohD , after drought stress i cheked the exporession of this gene in my plant.
i have two problem
1. I got High CT in my negetive husekeeping gene (GAPDH) but after check in agarose i didnt see any thing. I am worry about my final result after normalosation with this house keeping gene.
2. sometimes double melting curve in my specific gene only in one accession.
To point 1. How high was your Ct ? If the Ct is more than 6 cycles higher than the sample with the lowest expression, it should be fine. You are aware that higher Ct stands for lower expression, I assume ?
To point 2 what do you mean by accession ? How does this look on the gel ? Primer dimers perhaps ?