Specific Gene Silencing in Leptospira biflexa by RNA-Guided Catalytically Inactive Cas9 (dCas9)

Methods Mol Biol. 2020:2134:109-122. doi: 10.1007/978-1-0716-0459-5_10.

Abstract

Easy, practical, and affordable gene silencing techniques are constantly progressing, and genetic tools such as TALEs, RNAi, and CRISPR/Cas9 have emerged as new techniques for understanding the basic biology and virulence mechanisms of pathogenic organisms, including bacteria. Here, we describe one-step targeted gene silencing in Leptospira biflexa by using plasmids expressing catalytically inactive Streptococcus pyogenes Cas9 (dCas9) and a single-guide RNA (sgRNA) capable of pairing to the coding strand of a desired gene.

Keywords: Electroporation; Gene silencing; Leptospira; Single-guide RNA; dCas9.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • CRISPR-Associated Protein 9 / genetics*
  • CRISPR-Cas Systems / genetics
  • Gene Knockdown Techniques / methods*
  • Gene Silencing / physiology*
  • Leptospira / genetics*
  • Plasmids / genetics
  • RNA, Catalytic / genetics*
  • RNA, Guide, CRISPR-Cas Systems / genetics*
  • Streptococcus pyogenes / genetics

Substances

  • RNA, Catalytic
  • RNA, Guide, CRISPR-Cas Systems
  • CRISPR-Associated Protein 9

Supplementary concepts

  • Leptospira biflexa