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New to ChIP: bad Ct values


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#1 JustAnIntern

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Posted 30 June 2009 - 07:01 AM

I have extremely limited lab experience, and this is the first time that I've tried doing ChIP. I used the SABiosciences ChampionChIP kit and human leukocytes. When I did the qPCR, the Ct values for the target gene were generally higher than those of the negative control. Could this be a problem with the concentration of DNA? Also, I was wondering what is considered average for the Ct values. Most of mine fell between 25 and 35. I looked around online, and couldn't find any reviews of this kit, or anyone who has previously worked with this specific kit. Has anyone used this kit before and encountered the same problem? Any suggestions as to how to fix this problem would be greatly appreciated.

--JustAnIntern--

#2 cellcounter

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Posted 01 July 2009 - 05:11 PM

I have extremely limited lab experience, and this is the first time that I've tried doing ChIP. I used the SABiosciences ChampionChIP kit and human leukocytes. When I did the qPCR, the Ct values for the target gene were generally higher than those of the negative control. Could this be a problem with the concentration of DNA? Also, I was wondering what is considered average for the Ct values. Most of mine fell between 25 and 35. I looked around online, and couldn't find any reviews of this kit, or anyone who has previously worked with this specific kit. Has anyone used this kit before and encountered the same problem? Any suggestions as to how to fix this problem would be greatly appreciated.

--JustAnIntern--

Haven't used the kit, but I take anything outside of 15-30 as bad Ct value. Of course, this is too much simplification; there are way too many considerations to be made before you throw out any result. I can go into some details if you are so inclined but I would rather that you do some research into this. Looking up these protocols may help.




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