troubleshooting with site directed mutagenesis - (Jul/07/2008 )
Hi,
Am new to this field. I had cloned a gene (600 bp) for my work. I want to mutate one amino acid and tried SDM. After completing, I sent for sequencing for confirmation, but realised my primer sequence is repeated 6 times in the protein. Can anyone tell me the reason for this repeats??
Thank u in advance.
Some bacteria do weird things like recombination... if so, it's probable that repeats are consecuence of recombination... what bacteria stock are you using?
Am new to this field. I had cloned a gene (600 bp) for my work. I want to mutate one amino acid and tried SDM. After completing, I sent for sequencing for confirmation, but realised my primer sequence is repeated 6 times in the protein. Can anyone tell me the reason for this repeats??
Thank u in advance.
What's the primer sequences you used for SDM? Sufferd a bit from cases like this too (partial repeats of my primers, not complete ones) and it was due to the primer sequence (couldn't do too much about it besides introducing one or 2 silent mutations, which helped, but still had to check a lot more clones than usual to get a correct one).