Hello all,
I want to ask if you can proceed with under-sonicated chromatin by thawing on ice and sonicating extra cycles, will affect/compromise the chromatin quality laters with IP? I am doing double fixation protocol (DSG + FA) and sonicate using Bioruptor with 0.1% SDS final Conc. lysis buffer.
You can, but only if you did not perform the centrifugation and the spined pellet of debris is still present in the tube or preserved in the separate tube in the second case you need to mix the supernatant with the pellet resuspend by vortexing, re-sonicate, and then perform another spend down and collect supernatants. If the samples are from easy to obtain source (i.e. cell culture) I would restart the experiment, but if the samples are very valuable and difficult to reobtain it worth the try.
In my experience you can re-sonicate samples that are kept in sonciation buffer and not purified however this is likely to impact the IP efficiency, and may be better practice to simply start fresh with a new sample.
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