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Multiple-Gene Silencing Using Antisense RNAs in Escherichia coli

  • Nobutaka NakashimaEmail author
  • Shan Goh
  • Liam Good
  • Tomohiro Tamura
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 815)

Abstract

We have developed four expression vectors to express antisense RNAs (asRNAs) by which genes of interest are silenced in Escherichia coli. The vectors are all IPTG-inducible and co-transformable in any combination and target genes are silenced conditionally and concurrently. Furthermore, in order to improve silencing efficacy, the vectors are designed to express uniquely shaped antisense RNAs, named paired termini antisense RNAs (PTasRNAs). The vectors are useful for comprehensive investigation of gene function and are applicable even if the target genes are essential for cell growth. Here, we describe methods to construct PTasRNA-expressing vectors and to evaluate silencing efficacy.

Key words

Gene silencing Antisense RNA Escherichia coli Plasmid compatibility Co-transformation, Metabolic engineering Essential gene RNA silencing 

Notes

Acknowledgments

We thank the members of our research group for their assistance. This work was supported by Kato Memorial Bioscience Foundation.

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Nobutaka Nakashima
    • 1
    Email author
  • Shan Goh
    • 2
  • Liam Good
    • 2
  • Tomohiro Tamura
    • 3
  1. 1.Bioproduction Research InstituteNational Institute of Advanced Industrial Science and Technology (AIST)SapporoJapan
  2. 2.Department of Pathology and Infectious Diseases, Royal Veterinary CollegeUniversity of LondonLondonUK
  3. 3.Bioproduction Research InstituteNational Institute of Advanced Industrial Science and Technology (AIST)TsukubaJapan

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