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
- - - - -

[Overexpression] Without upstream transcription terminator


  • Please log in to reply
4 replies to this topic

#1 lactamase

lactamase

    member

  • Active Members
  • Pip
  • 17 posts
0
Neutral

Posted 09 May 2017 - 02:12 AM

Hi all,

 

I have been told that during the cloning of gene for protein overexpression, there should be a terminator placed upstream of a promoter and gene, if there is another gene (e.g. resistant marker) upstream to the target gene. Otherwise, the expression of the target gene will be affected.

 

However, is that real?


Edited by lactamase, 09 May 2017 - 02:13 AM.


#2 bob1

bob1

    Thelymitra pulchella

  • Global Moderators
  • PipPipPipPipPipPipPipPipPipPip
  • 6,400 posts
515
Excellent

Posted 09 May 2017 - 05:54 AM

Yes, it is real. You also need the terminator to ensure that the protein is correct when expressed - otherwise transcription just continues and you will end up with a large nonsensical protein expressed from the bases 3' of the intended sequence.



#3 lactamase

lactamase

    member

  • Active Members
  • Pip
  • 17 posts
0
Neutral

Posted 15 May 2017 - 11:31 PM

Thank you very much pulchella, biggrin.png biggrin.png 

 

However, how could the upstream promoter affect the downstream promoter if a terminator is not located in between? What is the mechanism of this?

 

Yes, it is real. You also need the terminator to ensure that the protein is correct when expressed - otherwise transcription just continues and you will end up with a large nonsensical protein expressed from the bases 3' of the intended sequence.



#4 bob1

bob1

    Thelymitra pulchella

  • Global Moderators
  • PipPipPipPipPipPipPipPipPipPip
  • 6,400 posts
515
Excellent

Posted 16 May 2017 - 10:04 AM

It's called transcriptional run-through - basically the transcriptional machinery will keep transcribing the bases beyond where you want (or should) have them terminate, meaning that you get nonsensical expression of the bases between the upstream and downstream genes. Without an internal ribosome entry site (IRES) this will prevent separate transcription of the downstream gene.



#5 lactamase

lactamase

    member

  • Active Members
  • Pip
  • 17 posts
0
Neutral

Posted 29 May 2017 - 11:04 AM

It's called transcriptional run-through - basically the transcriptional machinery will keep transcribing the bases beyond where you want (or should) have them terminate, meaning that you get nonsensical expression of the bases between the upstream and downstream genes. Without an internal ribosome entry site (IRES) this will prevent separate transcription of the downstream gene.

 

Thank you very much Bob, it is very much helpful smile.png smile.png 






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

©1999-2013 Protocol Online, All rights reserved.