PCR smear with CHIP - (Aug/23/2007 )
I have problem with Chip PCR. PCR result appears smear. Actually, I succeeded in doing CHIP experiment without any problem one year ago. However, when I started to do again recently following the same protocol I used before, I cannot get PCR product even in the sample with H3 antibody to pull down. H3 and primers worked well before. I doubted I did not have DNA in my sample, but running agarose gel test told me there were bunch of DNA there in the H3 pulling down sample with 0.35ug/ul after measurement by spectrophotometer. Can anyone help me out? Thanks.
-niw200-
QUOTE (niw200 @ Aug 23 2007, 03:55 PM)
I have problem with Chip PCR. PCR result appears smear. Actually, I succeeded in doing CHIP experiment without any problem one year ago. However, when I started to do again recently following the same protocol I used before, I cannot get PCR product even in the sample with H3 antibody to pull down. H3 and primers worked well before. I doubted I did not have DNA in my sample, but running agarose gel test told me there were bunch of DNA there in the H3 pulling down sample with 0.35ug/ul after measurement by spectrophotometer. Can anyone help me out? Thanks.
Are you using the exact same reagents you did a year ago? You might need new stuff...did your controls work?
-microbiobeast-
I tested my new stuff, looks fine. My input sample works great.
QUOTE (microbiobeast @ Aug 24 2007, 11:38 AM)
QUOTE (niw200 @ Aug 23 2007, 03:55 PM)
I have problem with Chip PCR. PCR result appears smear. Actually, I succeeded in doing CHIP experiment without any problem one year ago. However, when I started to do again recently following the same protocol I used before, I cannot get PCR product even in the sample with H3 antibody to pull down. H3 and primers worked well before. I doubted I did not have DNA in my sample, but running agarose gel test told me there were bunch of DNA there in the H3 pulling down sample with 0.35ug/ul after measurement by spectrophotometer. Can anyone help me out? Thanks.
Are you using the exact same reagents you did a year ago? You might need new stuff...did your controls work?
-niw200-