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Transcriptional Knockdown in Pneumococci Using CRISPR Interference

  • Morten KjosEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1968)

Abstract

Sequence-specific knockdown of gene expression using CRISPR interference (CRISPRi) has recently been developed for Streptococcus pneumoniae. By coexpression of a catalytically inactive Cas9-protein (dCas9) and a single guide RNA (sgRNA), CRISPRi can be used to knock down transcription of any gene of interest. Gene specificity is mediated by a 20 bp sequence on the sgRNA, and new genes can be targeted by replacing this 20 bp sequence. Here, a protocol is provided for design of sgRNAs and construction of CRIPSRi strains in S. pneumoniae, based on the vectors published by Liu et al. (Mol Syst Biol 13:931, 2017).

Key words

CRISPRi dCas9 Knockdown sgRNA Inverse PCR 

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Faculty of Chemistry, Biotechnology and Food ScienceNorwegian University of Life SciencesÅsNorway

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