Detection primer, detection kit and detection method of Leber's hereditary optic neuropathy (LHON) mitochondrial DNA gene mutations

Abstract

The invention relates to a detection primer, a detection kit and a detection method of Leber's hereditary optic neuropathy (LHON) mitochondrial DNA gene mutations. The invention, on the basis of Sanger sequencing principle, has the characteristics of being highly sensitive, stable and accurate; four most common pathogenic mutations of LHON mtDNA can be simultaneously detected, so that the problems of an existing LHON mitochondrial DNA mutation detection method, which is low in sensitivity, not strong enough in specificity, capable of causing pollution easily, high in expense and the like, can be solved; therefore, an accurate gene diagnosis method is provided for LHON; and the invention has important implications for genetic counseling and gene therapy of the disease (the LHON).

Description

[0001] Technical field: [0002] The invention belongs to the technical field of bioengineering, and in particular relates to a detection primer, a kit and a detection method for the mutation of the mitochondrial DNA gene of Leber's hereditary optic neuropathy. [0003] Background technique: [0004] Leber's hereditary optic neuropathy (LHON) is one of the most common mitochondrial genetic diseases and one of the most common causes of vision loss in young adults [Man PYW et al. J MedGenet, 2002]. More than 95% of patients are caused by three primary pathogenic mutations in mitochondrial DNA (mtDNA), including m.11778G>A in the MT-ND4 gene, m.14484T>C in the MT-ND6 gene and MT-ND1 gene m.3460G>A [Cerelli V et al. Prog Retin Eye Res, 2004]. The m.3635G>A mutation located in the MT-ND1 gene is the fourth common mutation in the Chinese population, and its mutation frequency is comparable to the m.3460G>A mutation [Jia XY et al. J Hum Genet, 2006]. Mutation detectio...

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Problems solved by technology

[0005] At present, on the one hand, there is no commercial LHON common mtDNA 4-site mutation detection kit on the market; on the other hand, the most common method for the genetic diagnosis of LHON has allele-specific PCR (AS-PCR) [CN1288254C, CN102146443A , CN101781683A], restriction fragment length polymorphism (RFLP) [CN1143117A, CN102747152A, CN104774927A, CN103173545A, CN104774926A, CN103173544A] and allele-specific fluorescent probe technology PCR [CN1721839A, CN1712543A common, detect single or 3] Mutation sites, these detection methods have the advantages of simplicity, rapidity, and low cost. However, due to the high frequency of mtDNA mutations in the population, mutations located in the vicinity of the primary mutation will affect the results, resulting in false positives or false negatives. In addition, There are also chip detection methods [CN101497925A, CN104805210A], which can detect more LHON-related mtDNA variations at the same time, and have the advantages of accurate and high-throughput, but this method is expensive

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