CRISPR/Cas12a-Assisted Ligation-Initiated Loop-Mediated Isothermal Amplification (CAL-LAMP) for Highly Specific Detection of microRNAs.
Anal Chem
; 93(22): 7942-7948, 2021 06 08.
Article
em En
| MEDLINE
| ID: mdl-34038095
ABSTRACT
Loop-mediated isothermal amplification (LAMP) has been increasingly applied in nucleic acid detection for clinical diagnosis and monitoring pathogenic microorganisms due to its isothermal nature and high sensitivity. However, the false-positive signal resulting from the non-specific amplification and the complexity of primer design are still technically challenging for wide applications. In this paper, we developed the CRISPR/Cas12a-assisted sequence-specific detection of LAMP products to eliminate the effect of non-specific amplification from primer dimers and spurious amplicons. Moreover, by designing a pair of target-specific stem-loop DNA probes, we greatly simplified the primer design for LAMP. The DNA probes could be ligated to form a double-stem-loop DNA template by the detected target, which initiated LAMP reaction and achieved one-nucleotide resolution due to the highly specific ligase reaction. Using microRNAs (miRNAs) as the model targets, the CRISPR/Cas12a-assisted ligation-initiated loop-mediated isothermal amplification (CAL-LAMP) can sensitively detect as low as 0.1 fM miRNAs with high specificity.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
MicroRNAs
/
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas
Idioma:
En
Ano de publicação:
2021
Tipo de documento:
Article
País de afiliação:
China