extraa bases before RE site - Reply urgent!! (Jun/01/2011 )
I have a basic doubt what if I dont keep any extra bases before the restriction site.. Would this create any problems in cloning of PCR products..
Please reply fast! As I am facing this problem.. but not sure whether it is because of this..
If you are digesting your pcr product before ligation, maybe. Some enzymes need few extra bases to 'land on'.
If you are cloning pcr ptoduct into simple vector as pGem t easy, no.
Also depends which polymerase you are using, some eat the ends of primers if you take your time seting up the reaction.