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Old 04-02-2019, 12:47 AM   #3
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Location: HK

Join Date: Mar 2019
Posts: 2

Hi, Femta. Thank you for your answer and suggestion first.
As I didn't find too much smeared DNA on the gel run after library preparation, library fragmentation may not be the case here. In combination with the low portion of Sequencing pores in MinKNOW report (not shown above), the most possible reason is the failure of adaptor ligation step. That is, I had enough DNA, but only a small part of them (especially the shorter part) was ligated to adaptors successfully, and only ligated DNA can be sequenced.
And I have asked for help from nanopore support and other nanopore users. They suggested that a longer ligation time under temperature lower than RM would increase ligation efficiency. I would try that in the next trial.
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