TNT T7 kit (Promega) with (Stratagene) SP6 pol

peter via methods%40net.bio.net (by peter.ianakiev from gmail.com)
Fri Feb 6 08:33:55 EST 2009


On Feb 6, 4:15 am, rompu... from gmail.com wrote:
> Hello,
> My lab has the TNT T7 Coupled Reticulucyte Lysate System (Promega).
> However the cDNA I want to transcribe/translate is in a plasmid under
> the control of Sp6 promoter. My lab has the Sp6 RNA polymerase from
> Stratagene. Therefore, has somebody try to add to the TNT mixture a
> different RNA polymerase (from a different company)? I have found some
> posts describing a buffert that can be used in that kit. However,
> these posts were related to T7 RNA polymerase.
>
> I would prefer to avoid subcloning the cDNA in a different plasmid and/
> or carrying out transcription/translation in two steps.
>
> Thanks
> Rom

I would not mess with the kit. Just make RNA separately with the SP6
pol , purify and use it with the reticulocyte extract.
my2c


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