Cloning large DNA fragments

Andy Cannons ACANNONS at DEAN.MED.USF.EDU
Tue Nov 23 17:09:06 EST 1993


Does anyone have some words of wisdom for us.  We are trying to 
subclone a 10kB genomic fragment from a cosmid clone into a 
plasmid.  However, we are experiencing alot of difficulties.  One 
problem is that even though the DNA fragment is gel purified some 
cosmid DNA must end up in our ligation mix and preferentially get 
transformed.  All clones we end up with appear not to be ligated into 
the vector we are using, bit recircularized cosmid.  Ideally we want a 
vector with 2 antibiotic resistance genes and MCS (Sac1 site if 
poss.).  Does one exist?  Anyother ideas to overcome our problems?

Andy Cannons




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