Nextera XT from ds cDNA

[kmkocot]

Hi all,

I made cDNA libaries using the Clontech SMART-Seq v4 ultra low RNA kit and made Nextera XT sequencing libraries from that cDNA. The bioanalyzer result (DNA HS) for the Nextera XT libraries is attached. Obviously Anamenia failed but I'm debating whether the others are OK for pooled sequencing. There is a really large variation in size range. Any input would be very welcome.

Thanks,
Kevin

[nucacidhunter]

Technically they can be sequenced. Size variation would be result of variation in input quantity for tagmentation and also bead:library volumes in final clean up. It will be difficult to get similar read number for the libraries. I would suggest to consult with your bioinformatician to see if they can account for unwanted size variation.

[kmkocot]

Thanks for the quick reply. For what it's worth, I measured concentration by Qubit and was pretty careful during all steps, especially the clean-up, so I was surprised to see this much variation.