Plasmid recombination

Priit [iso-8859-1] Jõers pjoers at ebc.ee
Wed Jan 17 09:03:26 EST 2001


    I have been currently plaqued by recombination of pRS series vectors
(Sikorski and Hieter 1989) while creating a genomic library in high-copy
vector. To be more precise, it`s pRS 425 (LEU). The plasmid appears to
lose significant amount of sequences (however, I haven´t determined,
which ones) in bacteria and is even more shorter that CEN/ARS-based
vectors. I used E. coli strain DH10B, should be especially suitable for
amplification of eukaryotic libraries. I have come across this problem
before with cloning, but then I could always purify extra 10 minipreps
and had a good chance to have at least one accurate clone. With library,
the problem can not be overlooked. My question to community is: have any
of you experienced same problems with pRS-series plasmids and if so,
what have you done to correct it.






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