gene design - restriction site addition (Apr/10/2009 )
i am designing a synthetic gene and i need to have Sal1and EcoRV restriction site at the 5' and 3' terminals.can i just insert the restriciton site of the enzyme on both ends and then design the whole gene..or do i need any additional base pairs in addition to these restriction sites on 5' and 3' ends....
You need additional bases. I would add at least 6, preferably 8 bases 5' of your cut site. I would suggest switching to a different enzyme than SalI, which has a very poor reputation for cutting PCR fragments.
phage434 on Apr 10 2009, 03:57 AM said:
XhoI and SalI form compatible overhangs