Hi everyone,
it's now several weeks that I have problems for the sequencing of mouse samples with the MinION device (from Oxford Nanopore Technologies). For that, I extracted gDNA from mouse tails (transgenic Parkinson's Disease mice) by using different Qiagen Kits (DNeasy® Blood & Tissue, QIAmp DNA mini kit). After an overnight dissociation of the tissue at 56°C, I extracted gDNA. Everything was fine and the ratios were good (260/280: 1.85; 260/230: 2.00) and the migration of the samples on a 0.75% agarose gel looked also fine with a big band over 15 kDa. From these samples, I prepared the library with the rapid sequencing kit (Reference SQK-RAD004) with 400 ng of gDNA. Finally, during the run, it appeared that the sequencing did not work properly since we had only 5000 reads in 2h. :-((( I decided to wash the flow cell and rechecked the number of pores. Since the Flow Cell could be reusable, I prepared a new library with the Lambda DNA from the Rapid Seq Kit. And everything worked perfectly well. So, it means that I have an issue with the samples that I prepared (even if I did the gDNA extraction twice). But I don't know why. Did somebody already have this problem? Do you have any idea/suggestions about how to fix this issue?
Thanks in advance. Tony
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