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ChIP-Seq for small number of cells

Genetics as it applies to evolution, molecular biology, and medical aspects.

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ChIP-Seq for small number of cells

Postby goldway » Tue Feb 22, 2011 6:35 pm

We are trying to do ChIP-seq using small number of cells (10,000-20,000 cells). We tried the method from Dr. Bradley Bernstein' s group (Adli et al., 2010 Nat Methods), but the data quality is not very good.

Some previous studies used WGA4 kit to amplify the ChIP DNA from small number of cells, then folowed by ChIP-Chip study. I wonder anyone tried the same method for ChIP -seq using illumina solexa sequencing? Any suggestion would be appreciated!
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Re: ChIP-Seq for small number of cells

Postby toniso » Tue Apr 05, 2011 10:57 pm

We also tried the method by Adli et al but similarly faced the problem of getting not really high quality data. I tried it for a transcription factor though. Perhaps it works with histone modifications but it is certainly much more challenging with transcription factors.
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Postby JackBean » Wed Apr 06, 2011 5:18 am

what about contacting the author and ask, whether you could come to his laboratory to see, how is it done exactly?
http://www.biolib.cz/en/main/

Cis or trans? That's what matters.
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Re: ChIP-Seq for small number of cells

Postby fizza9 » Thu Aug 18, 2011 11:54 am

Hi there,
I would like to perform a ChIP-seq on primary cells (FACS sorted). However, I can get only a limited number of cells per mouse (~3000/mouse). So I'd end up with 20,000 cells, is that enough for ChIP-seq? How about reproducibility? I read the paper from the Myers' lab, they used 50ng DNA and amplified it. 20,000 cells should give me ~200ng DNA, that might work. Has anyone experience with small cell numbers for ChIP-seq? Is there a protocol or article describing ChIP-seq on small number of sorted cells?
Thanks a lot!
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Re: ChIP-Seq for small number of cells

Postby Paul1987 » Tue Aug 23, 2011 9:33 am

"After you’ve got your hands on the isolated chromatin from your samples, the first critical step in the ChIP-Seq process is shearing it into little bite-sized fragments. This can be done in two ways:
Enzymatic Digestion
Sonication
In ChIP-Seq the fragment size is especially important in order to create good sequencing libraries. In standard ChIP assays you might shoot for fragment lengths anywhere from 200-1000 bp, but for ChIP-Seq it’s a bit shorter, 100-300 bp on the SOLiD platform for example. Pickle suggests optimizing your shearing conditions ahead of time before diving into an experiment."
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