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Expression from T7 promoter/DH5 Alpha cells

T7 promoter

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#1 Kumar@Reading

Kumar@Reading

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Posted 08 March 2013 - 03:38 PM

Hi, Everyone
I have cloned a Y.pestis gene into pET1.2 cloning vector. This vector contains T7 promoter as well. I just checked the protein expression in DH5a harbouring this pJET1.2+Y.Pestis gene and I found very high level of protein expression without any IPTG induction. I was not expecting the desired Y.pestis protein expression from this construct as E.coli DH5a cells dont have T7 RNA polymerase to start transcription from T7 promoter. The gene is in right orientation, confirmed by restriction digestion and sequencing. Could anyone help me please why I am getting expression in DH5a cells without any IPTG induction as well.
Many Thanks




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