Jump to content

  • Log in with Facebook Log in with Twitter Log in with Windows Live Log In with Google      Sign In   
  • Create Account

Submit your paper to J Biol Methods today!
Photo
- - - - -

pETduet transcription


  • Please log in to reply
No replies to this topic

#1 labtastic

labtastic

    Veteran

  • Active Members
  • PipPipPipPipPipPipPipPipPipPip
  • 195 posts
18
Good

Posted 28 December 2016 - 01:58 PM

When two proteins are expressed from a pETduet vector, how are the two genes transcribed?

 

I ask because the plasmid map indicates there are two T7 transcriptional start sites but only one terminator. To me this would indicate you'd get a mixture of "orf1+orf2" and "orf2 only" transcripts depending on where the T7 polymerase lays down (with no "orf1 only" transcripts).

 

I am considering moving this dual MCS from pETduet to the Invitrogen pBad vector (see page 9) by cutting pETduet with Nco/XhoI and pasting into same cut sites of pBad. This would preserve the flanking arabinose indicible transcriptional start and stops, the orf1 RBS from the pBAD vector and the orf2 RBS coming from the pETduet vector. For this to work though, the polymerase would have to transcribe both orf's as a single transcript. 

 

I suppose there would be no need for any of this if someone knew of a commercially available dual expression pBad vector. ;)


Edited by labtastic, 28 December 2016 - 02:03 PM.





Home - About - Terms of Service - Privacy - Contact Us

©1999-2013 Protocol Online, All rights reserved.