My first post/question. Trying and failing (4X now) to make PCR-free Illumina libraries with PCR amplicons (gel purified and sized 135-175bp), using KAPA PCR-free kit. My input is 20ng/ul based on Qbit = 1ug total. Using 15uM adapter = 0.68uM final concentration. qPCR tells me I'm getting 1nM properly adapted fragments. Should I double adapter concentration? Any clues why my efficiency is so poor?
Egel pic (with hard to read tapestation) attached
Egel pic (with hard to read tapestation) attached
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