The present invention provides a use of a Cas protein, and a method and a kit for detecting target nucleic acid molecules. The method for detecting target nucleic acid molecules comprises adding a guide RNA, a Cas12a, and a nucleic acid probe into a reaction system containing target nucleic acid molecules to be detected, and detecting it after the reaction is completed.

Disclosed herein are methods and compositions for detecting Bordetella pertussis and Bordetella parapertussis by detecting the presence of the IS481 and IS1001 genomic insertion sequences, respectively.

A cleavage-based real-time PCR assay method is provided. In general terms, the assay method includes subjecting a reaction mixture comprising a) PCR reagents for amplifying a nucleic acid target, and b) flap cleavage reagents for performing a flap cleavage assay on the amplified nucleic acid target to two sets of thermocycling conditions. No additional reagents are added to the reaction between said first and second sets of cycles and, in each cycle of the second set of cycles, cleavage of a flap probe is measured.

A cleavage-based real-time PCR assay method is provided. In general terms, the assay method includes subjecting a reaction mixture comprising a) PCR reagents for amplifying a nucleic acid target, and b) flap cleavage reagents for performing a flap cleavage assay on the amplified nucleic acid target to two sets of thermocycling conditions. No additional reagents are added to the reaction between said first and second sets of cycles and, in each cycle of the second set of cycles, cleavage of a flap probe is measured.

The present invention relates to the field of molecular biology detection, more particularly to a method for fluorescent quantitative PCR. The method includes: 1) mixing an upstream and downstream primer pair, a fluorescent probe, and a PCR amplification reagent; and 2) carrying out the fluorescent quantitative PCR, where the fluorescent probe has two quenching groups, in which a first quenching group is located at a 3′ end and a second quenching group is labeled on a T base and is 10-15 nt apart from the first quenching group. Using the method for fluorescent quantitative PCR, background signals in the fluorescent quantitative PCR can be reduced; furthermore, the sensitivity of PCR can be improved, the occurrence of false negatives in detection can be reduced, and the amplification efficiency of the fluorescent quantitative PCR can also be improved.

The present invention relates to the field of molecular biology detection, more particularly to a method for fluorescent quantitative PCR. The method includes: 1) mixing an upstream and downstream primer pair, a fluorescent probe, and a PCR amplification reagent; and 2) carrying out the fluorescent quantitative PCR, where the fluorescent probe has two quenching groups, in which a first quenching group is located at a 3′ end and a second quenching group is labeled on a T base and is 10-15 nt apart from the first quenching group. Using the method for fluorescent quantitative PCR, background signals in the fluorescent quantitative PCR can be reduced; furthermore, the sensitivity of PCR can be improved, the occurrence of false negatives in detection can be reduced, and the amplification efficiency of the fluorescent quantitative PCR can also be improved.

The present disclosure relates to a quencher having a quenching effect on a fluorescent material exhibiting luminescence characteristics at an excited energy level, and various uses thereof.

The present disclosure relates to a quencher having a quenching effect on a fluorescent material exhibiting luminescence characteristics at an excited energy level, and various uses thereof.

A method for detecting a target nucleic acid fragment in a sample, the method including a step of bringing the sample into contact with a gRNA, a Cas protein, and a substrate nucleic acid fragment, in which the Cas protein expresses nuclease activity after forming a complex with the gRNA and the target nucleic acid fragment, the substrate nucleic acid fragment is labeled with a fluorescent substance and a quenching substance, when the substrate nucleic acid fragment is cleaved by the nuclease activity so that the fluorescent substance is separated from the quenching substance, the fluorescent substance emits fluorescence due to excitation light, and the contact is performed in a reaction space having a volume of 10 aL to 100 pL so that when the target nucleic acid fragment is present in the sample, a tripartite complex is formed, the substrate nucleic acid fragment is cleaved, and the fluorescent substance is separated from the quenching substance; and a step of irradiating the fluorescent substance with the excitation light and detecting the fluorescence, in which detection of the fluorescence indicates that the target nucleic acid fragment is present in the sample.

A method for detecting a target nucleic acid fragment in a sample, the method including a step of bringing the sample into contact with a gRNA, a Cas protein, and a substrate nucleic acid fragment, in which the Cas protein expresses nuclease activity after forming a complex with the gRNA and the target nucleic acid fragment, the substrate nucleic acid fragment is labeled with a fluorescent substance and a quenching substance, when the substrate nucleic acid fragment is cleaved by the nuclease activity so that the fluorescent substance is separated from the quenching substance, the fluorescent substance emits fluorescence due to excitation light, and the contact is performed in a reaction space having a volume of 10 aL to 100 pL so that when the target nucleic acid fragment is present in the sample, a tripartite complex is formed, the substrate nucleic acid fragment is cleaved, and the fluorescent substance is separated from the quenching substance; and a step of irradiating the fluorescent substance with the excitation light and detecting the fluorescence, in which detection of the fluorescence indicates that the target nucleic acid fragment is present in the sample.