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Sports muscle injury related mitochondrial detection site, detection method and application

A technology for muscle damage and mitochondria, which is applied in biochemical equipment and methods, microbiological determination/inspection, DNA/RNA fragments, etc., can solve problems such as difficulties, PCR amplification bias, and mitochondrial sequence pollution, and achieve simple and convenient use , the effect of avoiding PCR amplification bias

Active Publication Date: 2022-03-04
GENERAL HOSPITAL OF PLA
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Moreover, the fragment length of most NUMTs is relatively short, ranging from tens to 200bp
Therefore, when short fragments are amplified or the amplified position sequence coincides with NUMTs, it is easy to cause bias in PCR amplification and contamination of mitochondrial sequences by NUMT sequences.
[0006] Therefore, it is currently difficult to judge the risk of muscle injury in athletes, especially before exercising. There is an urgent need for a method that can accurately and effectively judge the risk of muscle injury in athletes before training.

Method used

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  • Sports muscle injury related mitochondrial detection site, detection method and application
  • Sports muscle injury related mitochondrial detection site, detection method and application
  • Sports muscle injury related mitochondrial detection site, detection method and application

Examples

Experimental program
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Effect test

Embodiment 1

[0027] On the basis of the detection and screening of a large number of mutation sites in the early stage, according to the comparison between more than 30 people with rhabdomyolysis after exercise and the exercise tolerance group, it was found that among the reported mutations related to physical fitness, multiple gene loci There are significant differences in the genotyping of the points, so the Snapshot method is used to detect multiple sites, and these sites are detected simultaneously for genotyping, and finally the above 12 mutation sites with specific differences are screened out, so the It is combined into a detection site for sports muscle injury and applied in clinic. The specific mutation sites are shown in Table 1.

[0028] Table 1 12 mitochondrial pathogenic mutation sites

[0029] serial number mutation position mutation 1 3394 T>C 2 3421 G>A 3 4833 A>G 4 5814 T>C 5 8108 A>G 6 8363 G>A 7 9957 T>C ...

Embodiment 2

[0031] According to the mutation site detection panel provided in Example 1, two sets of long-fragment PCR primers and single-site one-step extension primer sequences were designed and detected. The specific sequences are shown in Table 2 and Table 3.

[0032] Table 2 Mitochondrial long-fragment amplification primer sequences

[0033]

[0034] Table 3 Single gene extension primer sequence

[0035]

[0036]

Embodiment 3

[0038] The long-segment primer and the single-site one-step extension primer in Example 2 were prepared as a kit. The kit includes long-segment PCR primer combination and single-site one-step extension primer mixture, and also includes: PrimeSTAR GXL DNA polymerase reaction mixture, deionized water, ExoSAP reaction enzyme, ddNTP with four kinds of fluorescent labels and extension The mixture of required synthetase and buffer system SNaPshot Multiplex, FastAP phosphodigestive enzyme and 10×FastAP Buffer.

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Abstract

The invention belongs to the technical field of detection of mitochondrial pathogenic sites related to motor muscle injury, and discloses a 12-site combination for detection of mitochondrial related to motor muscle injury, and amplification of mitochondrial genomes from two directions in a highly conserved region of mitochondrial, non-specific amplification of a mitochondrial DNA similar sequence carried by a nuclear genome is avoided, and two long fragments capable of covering a mitochondrial whole genome are obtained. The motor muscle injury related mitochondrial detection site provided by the invention is adopted as a detection object, and the kit provided by the invention is adopted, so that whether the corresponding site is mutated or not can be judged, if the corresponding site carries mitochondrial pathogenic base mutation, the existence of the susceptibility tendency of muscle injury during exercise training is prompted, and early warning is prompted.

Description

technical field [0001] The invention belongs to the technical field of sports muscle damage detection, and relates to a sports muscle damage related mitochondria detection site, detection method and application. Background technique [0002] In high-intensity, high-load exercise training, it may lead to excessive contraction or stretching of muscle fibers, resulting in injury-induced muscle cell necrosis, inflammatory cell infiltration, local inflammatory reactions, and body fatigue, muscle swelling, and soreness. Normal exercise fatigue is more difficult to recover from. Rhabdomyolysis occurs when muscle damage persists beyond the body's ability to recover, and the peripheral circulation presents substances released from the damaged muscle, causing tea-colored or even dark-colored urine. Release of muscle components into the blood can manifest as elevated levels of creatine kinase (CK), myoglobin, and α-actin, which can also be detected in urine Myoglobin. The level of c...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12N15/11
CPCC12Q1/6883C12Q2600/156
Inventor 吉栩杨广明姚咏明陈宁田亚平
Owner GENERAL HOSPITAL OF PLA
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