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Protocol for genomic recombineering in Yersinia ruckeri using CRISPR Cas12a coupled with the lambda Red system

Ejaz, Rooshanie N. ; Taylor, Nicholas M.I. and Steiner-Rebrova, Eva M. LU orcid (2024) In STAR Protocols 5(2).
Abstract
Genomic manipulation of Yersinia ruckeri, a pathogen of salmonid fish species, is essential for understanding bacterial physiology and virulence. Here, we present a protocol for genomic recombineering in Y. ruckeri, a species reluctant to standard genomic engineering, using CRISPR Cas12a coupled with the λ Red system. We describe steps for identifying protospacer guides, preparing repair template plasmids, and electroporating Yersinia cells with Cpf1 and protospacer plasmids with homologous arms. We then detail procedures for genome editing and plasmid curing.
Abstract (Swedish)
Genomic manipulation of Yersinia ruckeri, a pathogen of salmonid fish species, is essential for
understanding bacterial physiology and virulence. Here, we present a protocol for genomic
recombineering in Y. ruckeri, a species reluctant to standard genomic engineering, using CRISPR
Cas12a coupled with the l Red system. We describe steps for identifying protospacer guides,
preparing repair template plasmids, and electroporating Yersinia cells with Cpf1 and
protospacer plasmids with homologous arms. We then detail procedures for genome editing and
plasmid curing.
Please use this url to cite or link to this publication:
author
; and
publishing date
type
Contribution to journal
publication status
published
subject
in
STAR Protocols
volume
5
issue
2
article number
103014
publisher
Cell Press
external identifiers
  • pmid:38615317
  • scopus:85190088394
ISSN
2666-1667
DOI
10.1016/j.xpro.2024.103014
language
English
LU publication?
no
id
a81e9dd2-8831-445c-9f17-f54fa7e22727
date added to LUP
2024-06-19 13:09:49
date last changed
2024-06-20 08:25:07
@article{a81e9dd2-8831-445c-9f17-f54fa7e22727,
  abstract     = {{Genomic manipulation of Yersinia ruckeri, a pathogen of salmonid fish species, is essential for understanding bacterial physiology and virulence. Here, we present a protocol for genomic recombineering in Y. ruckeri, a species reluctant to standard genomic engineering, using CRISPR Cas12a coupled with the λ Red system. We describe steps for identifying protospacer guides, preparing repair template plasmids, and electroporating Yersinia cells with Cpf1 and protospacer plasmids with homologous arms. We then detail procedures for genome editing and plasmid curing.}},
  author       = {{Ejaz, Rooshanie N. and Taylor, Nicholas M.I. and Steiner-Rebrova, Eva M.}},
  issn         = {{2666-1667}},
  language     = {{eng}},
  number       = {{2}},
  publisher    = {{Cell Press}},
  series       = {{STAR Protocols}},
  title        = {{Protocol for genomic recombineering in Yersinia ruckeri using CRISPR Cas12a coupled with the lambda Red system}},
  url          = {{http://dx.doi.org/10.1016/j.xpro.2024.103014}},
  doi          = {{10.1016/j.xpro.2024.103014}},
  volume       = {{5}},
  year         = {{2024}},
}