Plasmid DNA isolation from yeast for transformation of E. coli.
Ohno, Kinji, M.D.
ohnok at mayo.edu
Fri Sep 17 14:57:07 EST 1999
I$B%f(Jm trying the glass-beads method to transfer a shuttle vector from yeast to
E. coli. I$B%f(Jm using CaCl2 method for bacterial transformation. I tried the gl
ass-beads methods by Hoffman (Curr Prot Mol Biol, Section 13.11.1, 1997) and
by Kaiser (Biotechniques 14: 552, 1993). I noticed that XL1-Blue does not wo
rk well for this purpose. Hoffman describes that HB101 and its derivative, M
C1061, would work fine, but these strains are endonuclease A-positive and wo
uld not work well for automated sequencing. EndA-positive strains are not re
commended for sequencing by Qiagen or Promega, and by a scientific paper (NA
R 21: 1677, 1993)
Does anybody have a good experience with EndA-negative strains, or do EndA-p
ositive strains work well for automated sequencing?
Kinji Ohno, MD. PhD.
Neurology Research, Mayo Clinic, Guggenheim 801B
200 1st ST SW, Rochester, Minnesota 55905
e-mail: ohnok at mayo.edu
phone: 507-284-5100 (office)
fax: 507-284-5831 (office)
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