Probe and kit for detecting point mutation of DNA and application

Abstract

The invention relates to a probe and kit for detecting point mutation of DNA and application. The probe is provided with an AP site used for being digested by endonuclease IV (Endo IV), and the AP site is used for dividing the probe into a first sequence and a second sequence; and the probe can form a first substrate and a second substrate through complementation of partial sequences of the probe with a mutant strand and a wild strand separately, the first substrate and the second substrate separately have first activity and second activity that the first substrate and the second substrate are digested by the endonuclease IV to form a free single strand with a second sequence, and the first activity is higher than the second activity. Due to the fact that the Endo IV has unique enzyme digestion activity on different truncated dsDNA structures such as the first substrate and the second substrate, a unique Endo IV probe is designed, the Endo IV probe and a target chain form the special dsDNA structure, the sequence dependency of the Endo IV is greatly lowered, the detection universality is widened, and the detection cost is reduced.

Description

technical field [0001] The invention relates to the technical field of single nucleotide mutation detection, in particular to probes, kits and applications for detecting DNA point mutations. Background technique [0002] Single nucleotide variation (SNV) or other types of various DNA point mutations are associated with a variety of human diseases, and various DNA point mutations have been recognized as biomarkers for clinical diagnosis and treatment of human cancer, such as TP53, EGFR and RAS gene mutations. However, mutant DNA (MT) often coexists with large amounts of wild-type DNA (WT) in clinical samples at low abundance levels (<0.1%). Currently, various methods for detecting DNA point mutations have been developed, including DNA sequencing, polymerase chain reaction (PCR), and nucleic acid hybridization. DNA sequencing (Sanger sequencing, next-generation sequencing, etc.) and PCR (allele-specific PCR, blocking PCR, droplet digital PCR, etc.) are the most commonly u...

AI Technical Summary

Problems solved by technology

However, the common problem of the probe-based DNA point mutation detection strategy is that the probe needs to be repeatedly optimized for experimental conditions; especially when detecting different SNVs, in order to determine the sensitivity and selectivity of the probe, it is often necessary to pass Optimization of reaction temperature, introduction of auxiliary strands and other methods to determine, which undoubtedly increases the cost of probe design

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