expression problem in BL21(DE3)

Kyle Legate legatek at mcmail.cis.mcmaster.ca
Tue Mar 26 10:09:58 EST 2002


Hi all, I have a problem I hope someone can help me with. I'm trying to
express two proteins in BL21(DE3), one on a high copy vector under Amp
selection and one on a pACYC vector under Cm selection. I get great
expression from the high copy vector but nothing from the low copy vector.
I tried plating cells transformed with only the Amp plasmid on Amp/Cm
plates (100ug/ml amp and 70ug/ml Cm) and noticed to my dismay that the
cells grow just fine! I was under the impression that only cells
harbouring pLys were supposed to be Cm resistant--am I wrong? I'm
beginning to wonder if our DE3 stocks became switched with our pLys
stocks, which are supposed to be stored in a different box--not good since
I won't be able to transform them with pACYC.
	Also, do BL21(DE3) harbour any intrinsic antibiotic resistance?
I'm starting to clone into a tet resistance vector but don't want to
continue if BL21 are resistant to tet.
	Thanks for your help.

... . . .  .  .  .    .    .    .     .     .     .      .      .      .
legatek at mcmaster.ca		Kyle Legate            legatek at hotmail.com

   Tower of Tongues:Thursday PM:10:30-11:30 EDT:http://cfmu.mcmaster.ca
    moon musick:ritual:IDM:experimental(electronica):minimalism:glitch
.      .      .     .     .     .    .    .    .   .   .   .  .  . . ...




More information about the Methods mailing list