Endofree Maxiprep and TOP10 cells

Frank O. Fackelmayer Frank.Fackelmayer at uni-konstanz.de
Sat Jul 15 14:20:54 EST 2000


Use CsCl to purify your plasmids. Nothing compares to the purity and
yield of this "old" standard method. In contrast to all the "newer"
stuff, CsCl is really independent of strain, growth conditions, and
plasmid copy number. And transfections with CsCl-DNA work every time,
with incredibly low cell death rates (<5% even with cells that are
sensitive to endotoxins).

Frank



Bertrand Collet wrote:


> Hi everybody,
> 
> I am having some problem of DNA quality with the Endofree Maxiprep
> Plasmid purification kit: A260/A280 ratio are never above 1.4-1.5 when
> this protocole is supposed to give ratios above 1.8. I tried to reduce
> the volume of the starting culture, use twice reagent for the alkaline
> lysis ... there is no way to improve the DNA quality. Are the TOP 10
> cells in cause (high carbohydrate content ...) ? Is it worth changing
> bacteria strain ? Is there anybody who had this problem with TOP10
> strain ?
> I do need top quality DNA for transfection purpose.
> 
> Thanks for your help,
> 
> Dr Bertrand Collet
> University of Aberdeen
> Department of Zoology
> Fish Immunology Group
> Tillydrone avenue
> Aberdeen AB24 2TZ
> Scotland
> Phone: (+44) (0) 1224 273 796
> Fax: (+44) (0) 1224 272 396






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