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I am in process of preparing libraries with 10 ng and 100 ng mRNA for RNA-Seq. I ran a BioA chip (DNA HS) following second strand synthesis and bead purification. However my BioA traces are flat? At this level of mRNA input would I expect to see a library peak following ds cDNA? I just wanted a quick QC before proceeding along the LC process. Any ideas are welcome!
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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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