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CHIP-seq problem with yeast gDNA!!!


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#1 lorenzo



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Posted 31 August 2015 - 01:24 PM

hi there,


I'm trying to optimize a RAD52  chip seq in S. cerevisiae. 

I'm growing 500ml at 0.5OD of cells, fix with 1% formal for 1h, pellet, bead beat the cells and then sonicate using a covaris (1m volume).(size 100-500bp)

after the whole CHIP protocol, I'm doing a bioanalizer to check the size of the IP and there is almost just a small fat band at 35bp!!


what I am doing wrong??!? 


doing some optimization of the sonication step I noticed that a huge 35bp band of RNA appears after sonication and I must say that after the IP I didnt RNase treat the sample nor use a qiagen column to purify the IP (I did phenol-chlor and ethanol precipitation.) but even if the small band at the end of the IP protocol is RNA that I can easily remove, I still dont have much DNA in the IP to sequence!! 


any suggestions??!?!




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