Im trying to create point mutation in domain of gene of interest using enzynomics EZ-MIX kit. I believe this kit follow quick-change site directed mutagenesis method; PCR amplification of forward-reserve complemantary primer with the introduction of mutation in the middle of primer, follow by dpn1 digestion.
I am able to get the colonies after transformation but when i sent two clones for sequencing, sequencing result turn out there is repeated region of whole mutagenesis primer in both clones. May i know if anyone had experience this and how to overcome this problem. Im thikning sending more clones for sequencing may help.Thanks!!