LacZ PCR Problems
Posted 25 July 2011 - 12:11 PM
Posted 25 July 2011 - 12:47 PM
LacZ R (oIMR0040): CGTGGCCTGATTCATTCC
LacZ F (oIMR3054): ATCCTCTGCATGGTCAGGTC
LacZ amplicon: 315 bp
If so, are you running a control PCR with it? The one Jackson laboratory has the following control PCR associated with this LacZ assay:
Internal Control F (oIMR8744): CAAATGTTGCTTGTCTGGTG
Internal Control R (oIMR8745): GTCAGTCGAGTGCACAGTTT
Positive Control amplicon: 210 bp
We've run this assay many times without any problem, which suggests to me that your problem is one of contamination (in your Taq as you explained or other reagents).
I assume you've already changed all your reagents so unfortunately I cannot really give you much advice since you seem to be doing everything you can to get your problem solved. I would also say that having to DNase treat your Taq is simply too much work for something as "straight forward" (when it works) as genotyping mice.
Have you considered sending your samples to be genotyped by a company? Nowadays you can get your mice genotyped quite cheaply by companies like Mouse Genotype. Just a thought.
Posted 29 December 2018 - 04:00 PM
Hi, I am new to this post site. I know this post is years ago but just curious "cookmacj", did you end up figuring out what the problem was and solving this issue? My coworker is now having this same issue. I would like to hear what you did to fix this. Thanks.