ChIP assay loading control
Posted 09 March 2005 - 09:23 PM
I think the first way is just the start matteral is same , but can not tell the process. The second way is start the same protein for IP , I do not know, somebody help me?
Posted 09 March 2005 - 09:56 PM
Yes, first use equal number of cells. You can do this by using duplicate treatment dishes, one dish for ChIP and the other solely for cell counting.
The second control is to set aside input DNA (1/100) after adding the ChIp dilution buffer but before adding ChIP antibody. When doing ChIP PCR, amplify the input DNA as well as ChIPed DNA. the intensity of the input DNA amplification will tell you the amount of starting DNA.
Another control for subsequent steps such as those multiple washings is to amplify some control gene. For example, if you are studying gene activation by histone acetylatio, you can amplify the G3PDH gene because its promoter is supposed to be acetylated. But if you are studying some repressive histone modifications, it is hard to control because the result is absense of amplicon.
Posted 10 March 2005 - 07:16 AM
Thanks a lot! wll
Posted 10 March 2005 - 09:08 AM
Thousands of thanks!
Posted 10 March 2005 - 09:28 AM