SNP (Single Nucleotide 0olymorphism) detection specific primer, liquid-phase chip and detection method of RYR1 (Ryanodine Receptors 1) gene

Abstract

The invention discloses an SNP (Single Nucleotide Polymorphism) detection specific primer, a liquid-phase chip and a detection method of an RYR1 (Ryanodine Receptors 1) gene. The liquid-phase chip comprises wild type and mutant type ASPE (Allele Specific Primer Extension) primer pairs respectively designed aiming at the mutational sites of each type, microballoons respectively coated with a specific anti-tag sequence and primers used for respectively amplifying target RYR1 gene sequences with G341R, R614C, T2206M and / or G2434R sites. The SNP detection liquid-phase chip of the RYR1 gene has very good signal-to-noise ratio, and a designed probe and the anti-tag sequences have no cross reactions basically. The designed ASPE primers have very good specificity and can accurately distinguish various types of mutational sites. The detection method has simple steps, and four SNP sites can finish being detected in one step, thus the operation is convenient; and moreover, various uncertain factors existing in the processes of multiple operations are avoided, thus the detection precision is greatly improved.

Description

technical field [0001] The invention belongs to the field of molecular biology, relates to medicine and biotechnology, in particular to a specific primer for RYR1 gene SNP detection, a liquid phase chip and a detection method. Background technique [0002] As a calcium ion channel protein, ryanodine receptor 1 (RYR1) plays a key role in the release of calcium ions from the terminal pool of muscle cells and the stimulation of muscle cell contraction. Mutations in RYR1 have been identified as central core disease (CCD), multiminicore disease (MmD), malignant hyperthermia (MH) and core / rod disease (Core / rod disease, CRD) is the main reason. [0003] Since zhang reported that RYR1 was the causative gene of CCD in 1993, so far, 44 RYR1 mutations have been reported to be associated with CCD, including 39 missense mutations and 5 deletion mutations. Studies have shown that most CCDs are caused by mutations in the RYR1 gene. RYR1 is also the causative gene of MmD, but so far onl...

AI Technical Summary

Problems solved by technology

[0006] At present, there are almost no products on the market to detect RYR1 gene polymorphisms at home and abroad, most of which are still in the stage of experimental research and have not yet been commercialized. The existing detection technologies are mainly based on PCR, such as direct sequencing, fluorescent quantitative PCR, PCR -Single-strand conformational polymorphism analysis (SSCP) detection, these technologies have the disadvantages of low sensitivity, easy sample contamination, and high false positive rate. At the same time, due to the limitation of detection throughput, they cannot meet the clinical needs

However, the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis technology can only detect one mutation at a time, which is time-consuming and laborious.

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