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GST-tagged protein purification. - inclusion body formation of my GST taggged protein. (Aug/17/2006 )

hello all.
I am expressing a GST tagged protein in BL21-DE3 strain of e. coli. My protein is 36 kda and iam getting a correct band on SDS-PAGE i.e 36+26. But the problem is that, iam not getting my protein in supernatant and it is coming in inclusion bodies blink.gif . Iam using sonication to lyse cells,which is 15 sec. pulse for 10 min at 15%..
what strategy i should follow to get the protein in solution, so that i can avoid denatured purification. rolleyes.gif



maybe you should try to lysate your cells using a french press, it always worked for me.
If you want to continue trying the sonication method, I would sonicate it untill you get a clear lysate.
I hope it will work!



you can also play around with your induction conditions, lower temperature and inductant concentration can help to solubilise proteins.