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vector auto ligation problem

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#1 mhtprc



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Posted 23 March 2005 - 08:08 AM

Hi guys, my subclone insert into pGEX4T2 vectors. I picked up some colonies and did mini pre of plasmids, the products gave me positive PCR test result, so I did not do double digestion. But expression result gave me only GST protein. Any advice? I wonder why the PCR test is not dependable.

#2 george@CASE



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Posted 23 March 2005 - 11:46 AM

how do you know that it is only the GST being expressed?

is your DNA fragment in frame with the GST? is there a premature stop codon?

I'd still the do the double digest on your minipreps just to see if it is consistent with your PCR result.

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