protein expression problem

jspollack at hotmail.com jspollack at hotmail.com
Wed Oct 16 17:44:52 EST 2002


I'm trying to overexpress a protein for purification using a His-tag system.  I initially cloned my gene into pQE80 from Qiagen, which places the His tag on the N-terminus.  My clone checks out via sequencing, but I'm getting no expression.  I then cloned the gene into pQE60, which puts the His tag on the C-terminus.  Again, my clone checks out by sequence, but I'm getting expression of a protein roughly twice the size I'm expecting.  My protein is nowhere to be seen.  The protein may or may not be toxic.  I really don't know.  Toxicity could explain the pQE80 problem, but I'm at a loss to explain how I'm getting expression of a 2-fold larger protein in pQE60.  Any suggestions on what to try?

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