Characterization and Engineering of a Novel Miniature Eubacterium siraeum CRISPR-Cas12f System.
ACS Synth Biol
; 13(7): 2115-2127, 2024 Jul 19.
Article
em En
| MEDLINE
| ID: mdl-38941613
ABSTRACT
Cas12f nucleases are one of the most compact genome editors, exhibiting promising potential for in vivo therapeutic applications. However, the availability of active Cas12f genome editors remains relatively limited in the field. Here, we report the characterization and engineering of a novel miniature Cas12f endonuclease from Eubacterium siraeum (EsCas12f1, 433 amino acids). We elucidate the specific Protospacer Adjacent Motifs preference and the detailed biochemical properties for DNA targeting and cleavage. By employing rational design strategies, we systematically optimize the guide RNA of EsCas12f1, converting the initially ineffective CRISPR-EsCas12f1 system into an efficient bacterial genome editor. Furthermore, we demonstrate the capacity of EsCas12f1 for in vitro nucleic-acid diagnostics. In summary, our results enrich the miniature CRISPR-Cas toolbox and pave the way for the application of EsCas12f1 for both genome editing and in vitro diagnostics.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Eubacterium
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Sistemas CRISPR-Cas
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Edição de Genes
Idioma:
En
Revista:
ACS Synth Biol
Ano de publicação:
2024
Tipo de documento:
Article
País de afiliação:
China
País de publicação:
EEUU
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ESTADOS UNIDOS
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ESTADOS UNIDOS DA AMERICA
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EUA
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UNITED STATES
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UNITED STATES OF AMERICA
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US
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USA