help? cDNA variability

[mcAstrocyte]

Hi all,

I am making my first 10x single nuclei libraries and ran two chromium chips with 4 samples in each chip.

While for each chip, every well had the same number of nuclei, however, I noticed that the first two samples in each case had low levels of cDNA (5 fold less than the other samples) when ran on the BioAnalyzer.

I didn't notice any wetting issues and all samples came out of the chip opaque and with 100ul, so I presume I am losing cDNA in the clean-up process. I am using a multichannel pipette for most stages in the protocol. Any ideas what might be causing me to lose cDNA in some wells but not others?

Thanks!

[nucacidhunter]

If you are using master mixes for workflow steps, the issue could be with position variability of thermocycler.