Problems in a fish genomic library construction

LUIS BEJARANO ARDURA lba at Galeon.uca.es
Thu Jun 20 09:44:43 EST 1996


Hi netters,

We want to make a fish genomic library in Lambda GEM 12 vector (Promega). 
The Lambda GEM 12 arms have been cut with BamHI restriction enzyme and we 
have eliminated the central stuffer (14 Kb) by electrophoresis in low 
melting agarose gel. The genomic DNA has been cut with Sau3aI restriction 
enzyme and dephosphorilated. Part of this genomic DNA has been 
selectioned by electrophoresis low melting agarose gel about 10 - 20 Kb 
size. We have tried all kind of combinations between Lambda GEM 12 BamHI 
arms and our genomic Sau3aI DNA in the ligation reaction. We have also 
tried several host strains like LE392, NM538 and KW251. What could we 
check to increased our very low number of recombinants?. Many Thanks.

 




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