Hi,
I am planning to do ChIRPseq which requires sonication of chromatin down to around a 100-500bp range. The protocol recommends using a Bioruptor and suggests that it may take up to 4 hours for sonication to that level. We have access to a Covaris M220 but not a Bioruptor so I wanted to try the sonication using the Covaris machine.
One critical point is that the aim of the protocol is to pull down RNA which is bound to the chromatin so any sonication process needs to minimize RNA damage. We would also like to examine any proteins pulled down (the process uses biotin labelled oligos which bind to the RNA) probably using mass spectrometry so need the proteins in reasonable condition also.
The initial cross linking is done using glutaraldehyde which I believe requires a longer sonication than formaldehyde treated cells.
Any advice regarding the use of Covaris vs Bioruptor or suggested alternative protocols would be greatly appreciated.
Thanks
Joshua
I am planning to do ChIRPseq which requires sonication of chromatin down to around a 100-500bp range. The protocol recommends using a Bioruptor and suggests that it may take up to 4 hours for sonication to that level. We have access to a Covaris M220 but not a Bioruptor so I wanted to try the sonication using the Covaris machine.
One critical point is that the aim of the protocol is to pull down RNA which is bound to the chromatin so any sonication process needs to minimize RNA damage. We would also like to examine any proteins pulled down (the process uses biotin labelled oligos which bind to the RNA) probably using mass spectrometry so need the proteins in reasonable condition also.
The initial cross linking is done using glutaraldehyde which I believe requires a longer sonication than formaldehyde treated cells.
Any advice regarding the use of Covaris vs Bioruptor or suggested alternative protocols would be greatly appreciated.
Thanks
Joshua
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