Programming the trans-cleavage Activity of CRISPR-Cas13a by Single-Strand DNA Blocker and Its Biosensing Application.
Anal Chem
; 94(9): 3987-3996, 2022 03 08.
Article
em En
| MEDLINE
| ID: mdl-35193353
ABSTRACT
The precise and controllable programming of the trans-cleavage activity of the CRISPR-Cas13a systems is significant but challenging for fabricating high-performance biosensing systems toward various kinds of biomolecule targets. In this work, we have demonstrated that under a critical low Mg2+ concentration, a simple and short single-stranded DNA (ssDNA) probe free of any modification can efficiently prevent the assembly of crRNA and LwaCas13a only by partially binding with the crRNA repeat region, thereby blocking the trans-cleavage activity of the LwaCas13a system. Furthermore, we have demonstrated that the blocked trans-cleavage activity of the LwaCas13a system can be recovered by various kinds of biologically important substances as long as they could specifically release the blocker DNA from the crRNA in a target-responsive manner, providing a facile route for the quantification of diverse biomarkers such as enzymes, antigens/proteins, and exosomes. To the best of our knowledge, this is reported for the first time that a simple ssDNA can be employed as the switch element to control the crRNA structure and regulate the trans-cleavage activity of Cas13a, which has enriched the CRISPR-Cas13a sensing toolbox and will greatly expand its application scope.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas
/
Sistemas CRISPR-Cas
Idioma:
En
Ano de publicação:
2022
Tipo de documento:
Article