Hello everyone,
I'm trying to optimize sonication for ChIP-exo for better reproducibility. I have never utilized the Covaris S220 system and cannot find a protocol for bacterial DNA shearing for ChIP. Here are my thoughts
I am shearing E. coli and am targeting 250 bp fragments. Before sonication, I have crosslinked with formaldehyde and lysed with lysozyme. The settings I am thinking of using are the following:
Duty Cycle: 10%
Intensity: 5
Cycles per Burst: 200
Time: 130 seconds
Following sonication, I plan on performing a phenol chloroform extraction followed by Nucleospin columns and running a Bioanalyzer to determine fragment sizes. The purpose of this experiment is to optimize the sonication settings to get more reproducible data.
Any comments, advice, or protocols will be greatly appreciated.
Thanks!
-M
I'm trying to optimize sonication for ChIP-exo for better reproducibility. I have never utilized the Covaris S220 system and cannot find a protocol for bacterial DNA shearing for ChIP. Here are my thoughts
I am shearing E. coli and am targeting 250 bp fragments. Before sonication, I have crosslinked with formaldehyde and lysed with lysozyme. The settings I am thinking of using are the following:
Duty Cycle: 10%
Intensity: 5
Cycles per Burst: 200
Time: 130 seconds
Following sonication, I plan on performing a phenol chloroform extraction followed by Nucleospin columns and running a Bioanalyzer to determine fragment sizes. The purpose of this experiment is to optimize the sonication settings to get more reproducible data.
Any comments, advice, or protocols will be greatly appreciated.
Thanks!
-M
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