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Multi-gene gateway clone design for expression of multiple heterologous genes in living cells: Eukaryotic clones containing two and three ORF multi-gene cassettes expressed from a single promoter

Paper ID Volume ID Publish Year Pages File Format Full-Text
24529 43524 2008 10 PDF Available
Title
Multi-gene gateway clone design for expression of multiple heterologous genes in living cells: Eukaryotic clones containing two and three ORF multi-gene cassettes expressed from a single promoter
Abstract

Two types of eukaryotic operon-type Expression clones were constructed using the Multisite Gateway system employing six types of att signals. These clones harbored a DNA cassette containing two heterologous ORFs (cDNAs) or three heterologous ORFs in tandem downstream of a single promoter. The most promoter-proximal ORF was translated via a Kozak signal and the downstream one or two ORF(s) were translated as directed by internal ribosome entry site(s) (IRES). These clones were observed to produce two or three different proteins at levels that depended on the activities of the translational initiation signals used. With the intention of modulating the expression level of the first ORF, the translational initiation signals including a Kozak sequence and 11 different IRESs were investigated for their efficiency using a single ORF. The translational activity of these signals varied within a 10-fold magnitude. Using these results, expression at pre-described relative levels was achieved from the optional IRES of the respective ORFs in the cassette. Controllable expression at desired levels of two different ORFs directed by optional IRESs on a bicistronic construct, transcribed from a single promoter, was demonstrated.

Keywords
Multisite Gateway cloning; Multi-gene Expression clone; Simultaneous introduction of multiple cDNAs; Eukaryotic operon; Controllable expression of transgenes
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Multi-gene gateway clone design for expression of multiple heterologous genes in living cells: Eukaryotic clones containing two and three ORF multi-gene cassettes expressed from a single promoter
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 136, Issues 3–4, 10 September 2008, Pages 103–112
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us