No plasmid from trasformed BL21(DE3)

Cornelius Krasel krasel at wpxx02.toxi.uni-wuerzburg.de
Sat Jan 10 11:02:13 EST 1998


Ke-Xue Huang <khuang at BIOC.RICE.EDU> wrote:
> you guy say, BL21(DE3) is difficult, Why?

First, the disclaimer: I've worked only with BL21(DE3)pLysS.

The competencies of this strain are lousy, compared to JM109 or DH5a. I
use the protocol of Chuang et al. (PNAS 1989) which gives routinely
1e08 colonies per mg DNA in the latter strains. In BL21 I get maybe
1e05. Maybe I'm doing something wrong with my ligations, but I usually
need at least 1e07 to get sensible results.

For obvious reasons, BL21(DE3)pLysS is unsuitable for minipreps (you'll
always get contaminating pLysS). I think, however, that BL21 is not endA,
therefore the quality of your DNA will always be worse than for endA
strains.

YMMV,
--Cornelius.

-- 
/* Cornelius Krasel, U Wuerzburg, Dept. of Pharmacology, Versbacher Str. 9 */
/* D-97078 Wuerzburg, Germany   email: phak004 at rzbox.uni-wuerzburg.de  SP4 */
/* "Science is the game we play with God to find out what His rules are."  */



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