Random Library Cloning - (Sep/29/2008 )
I am trying to generate a random library of fragment 10-20kb. I sheared it with a needle and got what I needed. But my problem is that I cannot repair it with T4 Pol because it will create a mutation in the polylinker region and screw up my blue:white colonies. Is there any way that I can clone it into a vector without repairing with T4 Pol?
Any help would be appreciated,
I have no idea why you couldn't use T4 DNA polymerase. If you performed random fragmentation using a needle, your polylinker would be gone anyway. Using T4 DNA polymerase just polishes the ends so you can ligate. The polylinker region is usually in the vector where you are cloning the fragments into. I am thinking you are not going to fragment the vector.