Afl II ligations... any suggestions?

Name fred at
Tue Mar 3 19:48:04 EST 1998

Dear netters-

I've repeatedly tried to do some cloning using an AflII site...and I've
tried a couple of obvious tricks to get any useful efficiency, but to no
avail.  Specifically, I have a 650 bp. EcoRI/Afl II fragment I'm trying to
get into the same sites of pSL1180  (superlinker from pharmacia).

Production of the fragments goes well, and I even have tried CIP-ing the
vector (just in case...)...I've tried ligation +/- PEG, high and low
amounts of DNA, at 12 degrees C and room temp., and high and low ligase
(400 and 2000 NEB units).  

And all I get are (very few) weirdo clones; that is, clones i can't
explain from fer instance, insertion of a dimer of my insert.  (and these
represent 3x more colonies than without insert!)

Any ideas?

Thanks in advance!!

Micky Krol

Inst. for Molec. Virology


em-ay-kay-are-oh-ell at

(translate the first part into letters to use this address)

More information about the Methods mailing list