Gene mutation detection kit and application thereof

A technology for detection kits and detection reagents, applied in the field of genetics, can solve problems such as unsuitable for large-scale promotion, high price, and large amount of data

Pending Publication Date: 2022-02-18
NANJING MATERNITY & CHILD HEALTH CARE HOSPITAL +1
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the preparation of next-generation sequencing samples is cumbersome, the amount of data generated is large, and the requirements for splicing and bioinformatics analysis of later data are relatively high, which is not suitable for large-scale promotion
In addition, despite the great development of next-generation sequencing technology, its price is still relatively high

Method used

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  • Gene mutation detection kit and application thereof
  • Gene mutation detection kit and application thereof
  • Gene mutation detection kit and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0132] The detection of embodiment 1 mutation site

[0133] SNaPshot technology was used to detect 224 hotspot mutations in genes related to hepatolenticular degeneration, phenylketonuria, tetrahydrobiopterin deficiency, and methylmalonic acidemia, including ATP7B, PAH, PTS, MUT, and MMAHC. The specific experimental steps are as follows:

[0134] 1) Take 1 μl of DNA sample for 1% agarose gel electrophoresis to check the quality of the sample and estimate the concentration, and dilute the sample to a working concentration of 5-10ng / μl according to the estimated concentration.

[0135] 2) PCR reaction

[0136] Reaction system (10μl):

[0137]

[0138] PCR cycling program:

[0139] 95°C 5min; 11×(94°C 20s, 65°C-0.5°C / cycle 40s, 72°C 1.5min); 24×(94°C 20s, 59°C 30s, 72°C 1.5min); 72°C 2min; 4°C keep warm .

[0140] 3) PCR product purification

[0141] Add 1U SAP enzyme and 1U Exonuclease I enzyme to 10μl PCR product, incubate at 37°C for 1 hour, then inactivate at 75°C fo...

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Abstract

The invention provides a kit for detecting ATP7B, PAH, PTS, MUT and MMACHC gene mutation and application of the kit. Specifically, 224 hot spot mutations of ATP7B, PAH, PTS, MUT and MMACHC genes are detected by using an SNaPshot technology. 51 pairs of PCR (Polymerase Chain Reaction) primers are designed for amplifying 51 100-500bp fragments containing 224 sites, and 224 extension primers adjacent to target sites are also designed for single base extension. The kit provided by the invention has very high accuracy, can quickly carry out gene diagnosis of related diseases, and provides effective reference for clinicians.

Description

technical field [0001] The invention relates to the field of genetics, in particular to a gene mutation detection kit and its application. Background technique [0002] Inherited metabolic diseases are diseases in which gene mutations lead to the deficiency of certain enzymes, receptors or carriers necessary for maintaining the normal metabolism of the body or the abnormal occurrence of proteins, resulting in corresponding pathological changes and clinical manifestations. Although the incidence of a single inherited metabolic disease is low, the overall incidence can reach 1 / 500 of live births. At present, the genetic diagnosis of genetic metabolic diseases is mainly based on next-generation sequencing technology. However, the sample preparation of next-generation sequencing is cumbersome, the amount of data generated is large, and the requirements for splicing and bioinformatics analysis of later data are relatively high, which is not suitable for large-scale promotion. I...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12Q1/6858C12N15/11
CPCC12Q1/6883C12Q1/6858C12Q2600/16C12Q2600/166C12Q2531/113C12Q2537/143C12Q2533/101C12Q2521/525C12Q2545/113
Inventor 许争峰胡平谭建新姜正文余锋
Owner NANJING MATERNITY & CHILD HEALTH CARE HOSPITAL
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