IRES, expression of several genes

Victor Levenson U10784 at UICVM
Sat Nov 18 15:54:57 EST 1995


In article <kenfuri-2111951532460001 at chuken04.shinshu-u.ac.jp> kenfuri at gipac.shinshu-u.ac.jp (Kenichi Furihata, MD) writes:
>From: kenfuri at gipac.shinshu-u.ac.jp (Kenichi Furihata, MD)
>Subject: IRES, expression of several genes
>Date: Tue, 21 Nov 1995 15:32:46 +0900

>I would like to express a protein complex consisting of three different
>subunits in one CHO cell line. I have heard of IRES (internal ribosomal
>recognition site?) with which we can subclone two different cDNAs in one
>expression vector and express the two corresponding proteins in one cell
>line. Could anyone let me know what is IRES or any better idea ?


   IRES (internal ribosomal ENTRY site) or CITE (cap-independent translation 
enhancer) - containing RNA fragment assumes a specific conformation which 
allows ribosomal entry. Several papers have been published on the use of this 
sequence for simultaneous expression of two (or more!!) proteins using one 
promoter with several genes "linked" by IRESs (I can find the references, if 
you want). IRES is used in pCITE series of plasmids made by Novagen (no 
affiliation).

   According to the literature it is an effective way to ensure simultaneous 
expression of 2 or more proteins in the same cells. Beware, though, that ATG 
codon of your gene has to be in-frame with the rest of IRES and at a certain 
distance from the core structure; e.g. with  pCITE2 it should be within NcoI 
site (ccatgg), the 1st cite in the polylinker. This limitation makes it rather 
cumbersome to clone genes under CITE.

If you are interested in details e-mail me.

Victor Levenson



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