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Our institute is having trouble completing these long runs. They seem to die towards the end of both reads, camera goes out of focus, etc. They are being run on an upgraded HS2000. FYI, the samples all passed on MiSeq 2x300. We are looking for slightly more yield meta, amp.Has anyone had similar issues, any success, any advice?
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I guess it depends on how good of success you're after. We had a 2x250 v2 rapid run finish this morning and the results were relatively good. There was decreased quality toward the end of each read, but this was due primarily to two tiles (I think they were the exact same ones for each of the reads), so it turned out that the vast majority of reads were quite good. I should note that there was a bigger issue with the end of read 2, where a large block of tiles had problems. The latter issue combined with the same tiles showing some quality problems in both read 1 and 2 would be compatible with a camera problem, since there was an obvious per-tile difference...thereby suggesting that the problem isn't due to the chemistry itself.
I can't give any advice, as I don't personally deal with the sequencer, but that's at least the sort of thing that we're getting at the moment. -
With an n=1 we had no problems. Got about 250M reads per lane.Comment
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Are you guys using a native 2500 V4 capable instrument, or an older upgraded 2000?Comment
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Our run was on an upgraded older 2000.Comment
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I hope the problem has been solved by now!
Anyway: With 2 native HS2500 we regularly get >300M cpf per flowcell and between 80 and 85% Q30 for amplicon data.Comment
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We've given up for the time being in favor of V1 rapids. Our instrument is an older one that we bought in 2011 and then upgraded to 2500. We're thinking that the camera/optics aren't capable of running 500 cycles?Comment
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Cancer survival rates have significantly increased in the last few decades in the United States, reaching a combined 70% 5-year survival rate by 2021. Behind this number, there are years of research to find new therapies, drug targets, and early detection methods. But there is one core challenge that keeps slowing down these advances, and it’s about drug resistance.
There is no single reason why many patients don’t respond to treatment as expected. Cancer is...Today, 05:17 AM -
Love this - good data definitely starts from good input, and poor input can only give relatively poor data. I particularly like the mention of Nanodrop/absorbance based methods for quantification. It's such a toss up if you'll get an accurate reading or what amounts to a randomly generated number, and a lot of library/sequencing related issues can be traced back to poor quant.07-01-2026, 11:43 AM
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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...
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