Entering edit mode
7.8 years ago
maxwellb
▴
10
As the title says, my lab mate and I are generating gene libraries for a single cell RNA-seq experiment. We're following SMRT-seq 2 protocol for reverse transcription. Our yield is within the optimal range after 20 PCR cycles (300pg/uL). Post-amplification fragment length distribution on the tape station (or bioanalyzer) should show a peak at ~2k bp. We're seeing a relatively broad distribution, with a peak at ~6k bp. I've attached an image of our gel with two representative lanes showing fragment distributions.
I've searched for troubleshooting advice on how to interpret such a result but can't find anything online. Has anyone had this problem? How did you interpret it? How did you fix it?
