Hey,

I have a 7kb plasmid that I need to amplify, but inside there is a 21bp sequence I need to change to my own sequence. I don't have comfortable restriction sites for this (tried and it's not working out). I tried using two large oligos that contain the 21bp sequence in the middle with an overlapping area to the plasmid on each side and then do a DPN digest (illustration). However, the amplification either doesn't work or it creates unplanned results. Which method would you recommend to go about this?