New Effective Method of Lactococcus Genome Editing Using Guide RNA-Directed Transposition.
Int J Mol Sci
; 23(22)2022 Nov 12.
Article
em En
| MEDLINE
| ID: mdl-36430465
ABSTRACT
Lactococcus lactis is an important industrial microorganism and a widely used model object for research in the field of lactic acid bacteria (LAB) biology. The development of new L. lactis and related LAB strains with improved properties, including phage-resistant strains for dairy fermentation, LAB-based vaccines or strains with altered genotypes for research purposes, are hindered by the lack of genome-editing tools that allow for the easy and straightforward incorporation of a significant amount of the novel genetic material, such as large genes or operons, into the chromosomes of these bacteria. We recently employed a suggested system based on the CRISPR-Cas-associated transposon for the editing of the L. lactis genome. After the in-depth redesign of the system, we were able to achieve the stable incorporation of the fragments that were sized up to 10 kbp into the L. lactis beta-galactosidase gene. The efficiency of editing under the optimized conditions were 2 × 10-4 and 4 × 10-5 for 1 kbp and 10 kbp, respectively, which are sufficient for fast and easy modifications if a positive selection marker can be used.
Palavras-chave
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Bacteriófagos
/
Lactococcus lactis
/
Lactobacillales
Idioma:
En
Ano de publicação:
2022
Tipo de documento:
Article