A CRISPR‐Cas9 tool to explore the genetics of Bacillus subtilis phages
| Autor: | V.T. Lutz, L.P. Baena Lozada, T. Schilling, N.M. Kühne, A.D. Furrer, K. Otte, Robert Hertel |
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| Rok vydání: | 2020 |
| Předmět: |
0106 biological sciences
Trans-activating crRNA Genetics 0303 health sciences 030306 microbiology Oligonucleotide Cas9 Mutagenesis (molecular biology technique) Bacillus Phages Bacillus subtilis Biology biology.organism_classification 01 natural sciences Applied Microbiology and Biotechnology Bacillus Phage 03 medical and health sciences Mutagenesis 010608 biotechnology CRISPR Guide RNA CRISPR-Cas Systems Genetic Engineering RNA Guide Kinetoplastida |
| Zdroj: | Letters in Applied Microbiology. 71:588-595 |
| ISSN: | 1472-765X 0266-8254 |
| DOI: | 10.1111/lam.13349 |
| Popis: | Here, we present pRH030, a new CRISPR-Cas9 tool for the genetic engineering of Bacillus phages and beyond. It is based on the Streptococcus pyogenes cas9 with its native constitutive promoter, tracrRNA, and a gRNA precursor. The constitutive expression of Cas9 was conducive to the inactivation of viral attackers and enhanced phage mutagenesis efficiency up to 100%. The gRNA precursor can be built up to an artificial CRISPR array with up to 5 spacers (target sequences) assembled from ordinary oligonucleotides and directly cloned into pRH030. Required time and resources remain comparable to a single gRNA cloning. These properties make pRH030 an attractive new system for the modification of Bacillus phages and qualify it for research beyond genetic construction. |
| Databáze: | OpenAIRE |
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