A kind of glucocerebrosidase gene detection kit and its detection method

A detection kit and cerebrosidase technology, applied in the field of genetic engineering, can solve the problems of complex operation, long time-consuming, low-throughput, etc., and achieve the effects of reducing false positives, avoiding subjectivity, and using less time

Active Publication Date: 2020-06-19
GUANGDONG HUAMEI ZHONGYUAN BIOLOGICAL SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] However, the current main technology for detecting GBA is gene sequencing, which has the disadvantages of low throughput, complicated operation, and long time consumption.

Method used

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  • A kind of glucocerebrosidase gene detection kit and its detection method
  • A kind of glucocerebrosidase gene detection kit and its detection method
  • A kind of glucocerebrosidase gene detection kit and its detection method

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Effect test

Embodiment 1

[0048] Embodiment 1: Composition and method of use of glucocerebrosidase gene detection kit

[0049] The detection kit includes Taqman probes for specifically recognizing 4 polymorphic sites on the glucocerebrosidase gene and primer pairs corresponding to the gene fragments where the polymorphic sites are located, specifically as shown in the table below:

[0050] Table 3. Composition of the detection kit

[0051]

[0052] The using method of described glucocerebrosidase gene detection kit:

[0053] 1. Extraction of sample DNA

[0054] According to the actual situation of the sample, choose the appropriate method to extract the sample DNA, common methods include magnetic bead method, anion exchange resin method and alkaline lysis method, etc.;

[0055] 2. Fluorescent quantitative PCR reaction

[0056] The amplification system of the fluorescent quantitative PCR reaction is as follows:

[0057] Table 4. Real-time quantitative PCR reaction system

[0058]

[0059] Th...

Embodiment 2

[0064] Embodiment 2: the establishment of standard atlas

[0065] According to the using method of embodiment 1, the homozygous standard substance and the heterozygous standard substance in the glucocerebrosidase gene detection kit are detected, obtain as follows figure 1 , 2 , the detection spectrum of the homozygous standard product shown in 4, 5 and such as image 3 and Image 6 The detection spectrum of the heterozygous standard substance.

Embodiment 3

[0066] Embodiment 3: No. 1 sample detection

[0067] According to the using method of embodiment 1, No. 1 sample is detected, obtain as follows Figure 7 and Figure 8 As shown in the graph, it can be seen that sample No. 1 is a homozygous wild type.

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Abstract

The invention provides a glucose cerebrosidase gene detection kit which is characterized in that the detection kit comprises a Taqman probe for specifically recognizing four polymorphic sites on a glucose cerebrosidase gene and a corresponding primer pair for amplifying gene segments where the polymorphic sites are; the four polymorphic sites are separately N188S, F213I, D409H and L444P. The Taqman probe comprises a wild probe and a mutant probe. The primer pair comprises an upstream primer and a downstream primer. Genotypes of the four polymorphic sites of the glucose cerebrosidase gene can be quickly and efficiently detected by applying the detection kit. The kit is a fast, simple, economical and efficient gene detection kit for glucose cerebrosidase deficiency disease.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, in particular to a glucocerebrosidase gene detection kit and a detection method thereof. Background technique [0002] Mutations in the gene encoding glucocerebrosidase (Glucocerebmsidas, hereinafter referred to as GBA) can cause a deficiency of the enzyme, resulting in the accumulation of glucocerebroside in the monocyte-macrophages of the liver, spleen, bone, and central nervous system Accumulation and disease, resulting in hepatosplenomegaly, skeletal deformities, growth retardation, anemia and other clinical manifestations. [0003] However, the current main technology for detecting GBA is gene sequencing, which has disadvantages such as low throughput, complicated operation, and long time consumption. [0004] Fluorescent quantitative PCR technology is a relatively mature nucleic acid detection method. Its advantages are: easy operation, less pollution due to closed-tube operat...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6883C12N15/11
CPCC12Q1/686C12Q1/6883C12Q2600/156C12Q2537/143C12Q2563/107C12Q2545/114C12Q2561/101
Inventor 郑文彦黄源坚王邦超杜蔚安高静彭百华周玉吴智锋
Owner GUANGDONG HUAMEI ZHONGYUAN BIOLOGICAL SCI & TECH
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