Detection probe and detection method for nucleic acid aim sequence

Abstract

The invention relates to a process of applying immune chromatography test paper to detect the target sequence of ribonucleotide. A detecting probe comprises identification sites of nicked endonuclease, the 3' portion of the probe is marked with biotin, and the 5'portion of the probe is marked with SMD (sulfametoxydiazine). The probe forms an integral reaction probe through incubating with magnetic beads which are marked with streptavidin. The probe is nicked into two sections by enzyme sites on the nicked endonuclease identification probe after the hybridization of the integral probe and the target sequence of ribonucleotide, which reduces the stability of the combination between the integral probe and the target sequence and separates with target sequence. The target sequence can be hybridized with another integral probe and be nicked by nick endonuclease. Therefore, a magnification mode of detection signals is formed in the reaction system. The section of the probe with magnetic beads is absorbed to deposit, and the section of the probe with SMD presents on a free state in solution after reaction solution being acted with magnetic field. The probe with SMD can be detected in detecting solution through using the immune chromatography test paper so as to reach the effect of detecting the target sequence. The process is simple, fast and economical, which is suitable to detect the target sequence of ribonucleotide in large-scale field screening.

Description

Technical field: [0001] The invention relates to the field of molecular biology, in particular to a method for detecting nucleic acid target sequences by using immunochromatographic test paper. More specifically, it is a method involving the detection of target sequences using restriction endonucleases, modified oligonucleotide probes and immunochromatographic test strips. Background technique: [0002] Fluorescent RT-PCR technology was developed in the late 1990s. This technology puts fluorescein-labeled probes and primers together and reacts in a fluorescent PCR instrument. The computer monitors the entire reaction in real time to avoid cross-contamination. It has improved the detection sensitivity and has been successfully used in the detection of human hepatitis C virus, HIV, etc., but this kind of RT-PCR-based detection method requires an expensive PCR machine for the reaction, which takes a long time and generally requires The response time of 1-2 hours is not conduci...

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Problems solved by technology

[0011] ①The realization of these methods requires the cooperation of various enzymes, the reaction system is complex, and it is not easy to optimize the reaction conditions, and most of the enzymes or reagents used in these methods are expensive, such as: helicase in the HAD reaction, Q- Qβ replicase in β reaction, Φ29 DNA polymerase in RCA reaction, etc.

[0013] ③ The RCA reaction is the most sensitive among all isothermal amplifications, but its reaction process is complicated, and there are many times in the process of changing the reaction system

[0014] ④ The later detection of isothermal amplification still cannot get rid of the limitation of the instrument, which makes the application of this kind of method very limited, and it is not convenient to carry out rapid screening in public places and remote areas

[0015] It is precisely because of the above shortcomings that isothermal amplification has not been used on a large scale at this stage.

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