Problems in cloning with LambdaGEM-12
m.l.jones at bham.ac.uk
Fri Nov 4 05:31:58 EST 1994
LambdaGEM-12 Xho I half-site arms cloning system.
In this system lambdaGEM-12 DNA has been digested with Xho I, partially
filled in with dTTP and dCTP, and dephosphorylated. This allows it to
ligate specifically with Mbo I- or Sau 3A I-digested genomic DNA which has
in turn been partially filled in with dATP and dGTP.
Using this lambdaGEM-12 cloning system (from Promega) I have been
obtaining unusually high levels of background, where vector DNA has been
ligated and packaged in the absence of insert genomic DNA. These were
20,000 pfu/microgram of vector DNA. The lot numbers of the kits used were
188802 and 3242023. I was also using Promega's packagene extracts.
Has anybody else had similar problems? Any advice would be greatly
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