Hello!
I am doing a CRISPRi-screening. I prepared sequencing library by PCR amplification using a i5 common primer and i7 primers with single TruSeq index, and performed a paired sequencing on NovaSeq 6000. Unfortunately, the sequencing quality for R2 was very poor (as shown in the attached image) while R1 was normal. Anyone knows what might cause this phenomenon?
I'm sorry to hear that you're experiencing sequencing issues with your CRISPRi-screening library. The fact that only one of the paired-end reads (R2) is affected suggests that the problem is likely related to issues specific to that read or the sequencing chemistry.
Here are some potential reasons for poor sequencing quality in R2:
If the issue persists after checking these factors, it may be helpful to consult with the sequencing facility or the manufacturer's support team for further assistance. Additionally, bioinformatics tools and quality control metrics can be used to assess and filter out low-quality data during the analysis phase.
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