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Stable and enhanced gene expression in Clostridium acetobutylicum using synthetic untranslated regions with a stem-loop

Paper ID Volume ID Publish Year Pages File Format Full-Text
22644 43368 2016 4 PDF Available
Title
Stable and enhanced gene expression in Clostridium acetobutylicum using synthetic untranslated regions with a stem-loop
Abstract

•Short single-stranded UTR decreased gene expression in Clostridium acetobutylicum.•Addition of a stem-loop at the 5′ end of mRNA increased mRNA level.•Addition of a stem-loop at the 5′ end of mRNA increased protein expression level.

Gene overexpression is one of the most basic strategies in metabolic engineering, but the factors determining gene expression levels have been poorly studied in Clostridium species. In this study, we found that a short single-stranded 5′ untranslated region (UTR) sequence led to decreased gene expression in Clostridium acetobutylicum. Using an in vitro enzyme assay and reverse transcription-quantitative PCR, we found that addition of a small stem-loop at the 5′ end of mRNA increased mRNA levels and thereby protein expression levels up to 4.6-fold, possibly protecting mRNA from exonuclease attack. Gene-expression levels were apparently independent of the stability of the added stem-loop; the existence of a stem-loop itself appears to be more important. Our results indicate that efficient expression cassettes can be designed by taking the 5′ UTR into consideration, as the expression levels can vary even though the same promoter and RBS are used. These findings will be useful for developing a more reliable gene expression system for metabolic engineering of Clostridium strains.

Keywords
Clostridium acetobutylicum; Gene overexpression; 5′ untranslated region
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Stable and enhanced gene expression in Clostridium acetobutylicum using synthetic untranslated regions with a stem-loop
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 230, 20 July 2016, Pages 40–43
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