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Does anyone have any suggestions about how to filter out the mis-called variants on Nextseq? Also, has anyone used the v2 kits? Is the fidelity significantly improved?
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We are about to run a test on the NextSeq v2 chemistry, so I'll probably have some data in a week or two. -
Thank you. We are anxious to see if we can improve the reliability of our Nextseq data. Our group at the National Cancer Institute is attempting to identify mutated gene products expressed in cancer cells that can be recognized by patient immune cells that will allow us to develop effective cancer therapies. We purchased a Nextseq because we needed to get our data quickly in order to develop treatments for patients with advanced cancer, and we have already identified many targets using this approach. Enhancing the accuracy of the variant calls would make this process simpler, as it would help to eliminate many of the false positives. Some of these are easy to identify as they appear in repeated tumor samples - I presume these are regions of the genome that are difficult to sequence. We also merge whole exome with RNAseq, which helps to validate the calls.Comment
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I’m not a sequencing expert. I’m a purification scientist who uses NGS to evaluate workflows my group develops. With this perspective, we think about the sample first and the NGS workflow second. The sequencer is an exceptionally honest reporter, but it can only report on what you give it, so whether you get clean, interpretable data from an NGS workflow is largely determined before you begin.
Here are nine questions we think about, in roughly the order they matter, before...06-18-2026, 07:11 AM -
Data variability is still an issue in sequencing technologies despite the advances in reproducibility and accuracy of these platforms. But the problem does not originate in the sequencing itself, but in the previous steps, before the sample reaches the sequencer.
The first step is collection, followed by preservation and sample preparation for analysis. Most scientists overlook those steps, but not being careful might just be skewing the experiment’s results.
...06-02-2026, 10:05 AM
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