Competent cell - How good does it need to be? (Jul/07/2005 )

I've been having problem with cloning a 3.2kb fragment into a 5.5kb vector. I tested the efficiency of my competent cell recently and found that it's 3x10(to the power of 6) per microgram of DNA. Is this efficiency too low and affecting my cloning?

Hi,

That number looks acceptable to me for usual cloning.
I recently checked myself competent cells attached to
TOPO cloning kit and it was 5x10 (to the power of 7) per
microgram.

If you don't have any colonies, I'd also recommend that
you check ligation by running a small portion of the reaction
mixture on gel.

I use subcloning efficiency DH5 alphas from Invitrogen and they claim approx 1exp6/ug plasmid and they work just fine for me.

Hi Slab and pBluescript,

Does your competent cells work well even when you transform larger plasmids i.e. > 8kb?