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ChIP Kick

Member Since 12 Dec 2012
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Topics I've Started

Why there are blobs in my sheared DNA bands?

07 January 2013 - 10:29 AM

Hi, I have some problems with my sheared DNA now. Hope you can help me! Attached are two images for the same DNA gel. Image were taken at different time. One is at 1:00 pm and another is at 3:11pm.

It seems like that I get some smeared DNA. But I do not know why there are blobs at the bottom of the band. I treated samples with proteinase K, incubate at 58 degree Celsius, overnight. Then did RNase A treatment at 37degree Celsius for 1 hour. After that, run a agarose gel at 53 voltage. Most sheared DNA bands in web don't have blobs. Could you give me some suggestions about that?

Thanks a lot!

Used all ChIP lysis buffer

04 January 2013 - 12:51 PM

Hi,  I tried many times but I still cannot get a good sonication condition for my exp. Now I used all my lysis buffer which I bought from Millipore(EZ-Magna ChIP™ A Chromatin Immunoprecipitation Kit). I checked their web but no recipe for it.There are so many different recipes for lysis buffer. I do not know which one is good for my exp.

So is there someone who could give me suggestions to make my own cell lysis buffer and nuclear lysis buffer?

Thanks a lot!

Sonication's fault or gel's fault? Help please!

12 December 2012 - 12:51 PM

Hi guys,

I have some problems with my ChIP assay. Hope you can give me some suggestions.

I sheared some formaldehyde cross-linked tissues samples. Then, treated with RNase A at 37C for 30min. After adding protinase K, reversed crosslinking at 62C for 2hours. Then run a 1% agarose gel with 85voltage.

Here are the problems. I do not know why there are red lines(in attached file) in my gel when running? In gel images, why I do not have smeared DNA?

My sonicator is Qsonics Q125. Sonication with 30% power, 15sec on, 45sec off, 10cycyles.

Is there someone can help me? Thank you very much!

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