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A recent illumina run analysis showed that the majority of our sequences were contaminated with 16S rRNA. Prior to dscDNA we used a rRNA depletion method using the RiboZero Bacteria Magnetic kit. We ran our samples on a Pico Chip and saw our sample had lost the peaks from 16S and 23S so the depletion step works. If contamination occurred it must have been either during dscDNA construction or prior to the illumina run. Could you help me determine which step might have introduced contaminants? In my attachment you will see the dscDNA run on a high sensitivity chip. Does this look fine? Please ignore the empty lanes.
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Hi vanillasky,
In my experience, you should see a one and only smooth band in your dscDNA preparation when running a DNA HS chip… I can't see that in the file you attached. So I do not recommend you to proceed with sequencing for this sample.
I also think that after RIboZero treatment, running a NanoChip is enough and better than the Pico one. -
Thank you for the reply. Unfortunately, I did end up sequencing and found that there were contaminants in there. I was never able to get this particular sample to work with the library preparation kit I used. I have talked to Agilent tech support in the past and they recommended the pico chip for mRNA analysis (which was my target). They told me the nanochip is good for quantitation vs the picochip. Also, the concentrations of my sample are so low that working with the picochip is better. Anyway, thank you for your post, I will keep this in mind if I ever work on this again.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...Today, 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 -
With the launch of new single-cell sequencing platforms in 2026, the field stands at an exciting inflection point. This article surveys the most impactful advances in the field and discusses how they’re reshaping research in cancer, immunology, and beyond.
Introduction
Single-cell sequencing technologies have undergone remarkable advances over the past decade, transitioning from low-throughput experimental approaches to highly scalable platforms capable of...05-22-2026, 06:42 AM
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