Ran qPCR on DNased withering syndrome RNA to verify there’s no residual gDNA, prior to performing reverse transciption.
Master mix calcs are here: 20150318 – qPCR WSN DNased RNA Test
Used 1μL of template. This was based on using 17.5μL of DNased RNA in the reverse transcription reaction. Each sample was ~20ng/μL which results in ~350ng DNased RNA in a 25μL reverse transcription reaction (350ng/25μL = 14ng/μL). So, using 1μL in the qPCR reaction roughly approximates the amount of DNased RNA one will encounter when running qPCRs with the cDNA.
Positive control was the 3e6 p18RK7 standard curve sample from 20120731.
All samples were run in duplicate.
See qPCR Report (see Results) for plate layout, cycling params, etc.
The only reaction that amplified was the positive control. The DNased RNA does not have any amplifiable gDNA. Will proceed to reverse transcription.