screening lb agar plate for bacteria expressing a protein
Wolfgang.Schechinger at med.uni-tuebingen.de
Fri Mar 17 13:37:18 EST 2000
currently I try to get a cDNA into an expression vector. (a
modified pMAL-c2x from NEB). The cDNA is coding for His6 and a myc
epitopes and for GFP. For experts: malE is not functional anymore.
Since I got too many colonies to perform minipreps and RE digests on
every single one, I thought I would plate a replica on IPTG-plates
and check for protein expression first. Then, I would transfer the
bacteria as replica on a nitrocellulose membrane and make an ECL
on the blot with e.g. anti-myc.
Does a protocol exist for that procedure? The main problem probably
is how tho crack the bacteria and glue their proteins to the
membrane. Is there something easier (like flashing with blue light on
the plate and see the expressing bugs glow green)?
All input is welcome!
This message is encrypted. Use your brain to decode it.
Dr. Wolfgang Schechinger, Dept. of Pathobiochemistry
University of Tuebingen, Germany
email: wolfgang.schechinger at med.uni-tuebingen.de
usual disclaimers apply
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