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Hi, saw some library kits using sequential adatation process, first to add adapter to the 5 ' prime and then 3'prime, just wondering how it works, many thanks in advance!
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Hello,
This kind of protocol is typical of smallRNAseq.
It uses a truncated T4 RNA ligase 2 and a modified 3' adapter.
Trunc. T4 RNA ligase is unable to adenylate the 5' end of its substrate so it uses the modification of the 3' primer (rApp) to perform this step.
The second ligation uses a classical RNA ligase.
I wasn't aware of such protocols used for RNAseq... -
Thanks!
Swift Bio for example using this strategy to build their Libraries, for DNAseq and RNAseq. It should be different for DNA and RNA application, right?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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