RNA degradation after lysis in single cell droplet sequencing procedures?

0880484929 · 10-30-2018, 12:32 PM

Our lab has built a drop-seq setup. Going over the protocol, I see that the cells are lysed in a solution of sarkosyl, Ficoll-400, tris, EDTA, and DTT. I'm not intimately familiar with the biochemstry of this lysis buffer. I'm wondering if it is effective at inactivating endogenous RNAses? I haven't found anyone online that has discussed degradation of RNA at this stage. But the pumping can take quite a while, depending on the scale of the experiment (at least 15 minutes per sample). I am just curious if this is something I should or shouldn't be concerned about.

10-30-2018, 12:32 PM	#1
0880484929 Junior Member Location: USA Join Date: May 2017 Posts: 2	RNA degradation after lysis in single cell droplet sequencing procedures? Our lab has built a drop-seq setup. Going over the protocol, I see that the cells are lysed in a solution of sarkosyl, Ficoll-400, tris, EDTA, and DTT. I'm not intimately familiar with the biochemstry of this lysis buffer. I'm wondering if it is effective at inactivating endogenous RNAses? I haven't found anyone online that has discussed degradation of RNA at this stage. But the pumping can take quite a while, depending on the scale of the experiment (at least 15 minutes per sample). I am just curious if this is something I should or shouldn't be concerned about.