Typing optimization method for SNP loci of rheumatoid disease genes
A gene locus and rheumatoid technology, applied in the field of gene identification, can solve the problems of high cost, harsh experimental conditions, and long time consumption
Pending Publication Date: 2021-03-09
广东瑞昊生物技术有限公司
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Problems solved by technology
However, the above methods have certain disadvantages: SSCP can only use non-denatured glue, and the experimental conditions are harsh, requiring room temperature conditions, which are difficult to achieve in general laboratories; the experimental process of CAPs is cumbersome, and an experimental procedure takes a long time; MassARRAY mass spectrometry The cost of TaqMan probe method and other chip methods is quite high, and it is difficult for ordinary laboratories to afford
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Embodiment 1
[0084] A method for optimizing the SNP site typing of rheumatoid disease genes, such as figure 1 shown, including the following steps:
[0085] 1) Extract the genomic DNA in the sample to obtain the DNA extraction solution;
[0086] 2) Take 1 μl DNA extract obtained in step 1) for PCR amplification, add 1 μl PCR buffer solution, 3.0mmolMg 2+ Solution, 0.3mmol dNTP, 1U DNA polymerase (from Qiagen Inc.) and 1μl PCR primers were mixed for multiple PCR reactions to obtain PCR amplification products; the amplification conditions were: 95°C, 5min pre-denaturation; then sequentially at 94°C, 11 cycles of 20s, 65°C, 40s, 72°C, 1.5min; then 20 cycles at 94°C, 20s, 59°C, 30s, 72°C, 1.5min; then keep at 72°C for 2min; finally Store at 4°C.
[0087] PCR primers consist of 16 SNP gene loci, and the rs numbers of the 16 SNP gene loci in the SNP database in NCBI are: rs6682654, rs3766379, rs143383, rs7639618, rs10865331, rs13202464, rs17095830, rs4552569, rs2273061, rs2273061, rs13182402 ...
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The invention discloses a typing optimization method for SNP loci of rheumatoid disease genes. The method comprises the following steps: 1) extracting DNA; 2) performing PCR amplification on the DNA;3) purifying a PCR amplification product; 4) extending the purified PCR amplification product; 5) purifying an extension product; 6) performing loading to a sequencer; and 7) analyzing fluorescence labeling and length information of the extension product. Based on an SNaPshot technology, PCR amplification primers and extension primers, with different lengths are designed for different SNP loci toachieve typing of multiple SNPs in one reaction system, and typing can be performed at 16 loci at the same time. Due to the fact that the method adopts four-color fluorescence labeling, various SNP types can be classified, and besides, insertion and deletion can be analyzed. After the genotyping of a rheumatoid disease patient is obtained through analysis, prevention before diseases of the user and symptomatic medicine application of a doctor after the disease occurs are facilitated.
Description
technical field [0001] The invention relates to the technical field of gene identification, in particular to a method for optimizing the typing of SNP sites of rheumatoid disease genes. Background technique [0002] Rheumatoid disease is a disease that has slowly entered people's field of vision in recent years. The incidence of this disease is increasing year by year. At present, the incidence of rheumatic diseases in my country is about 30%, while the incidence of gout caused by rheumatism is rising rapidly. , has risen from 3% 10 years ago to about 10% at present, so people have to pay attention to it. The incidence of this disease will always be concentrated in middle-aged and elderly people aged 50-60, and the incidence rate of women will be slightly higher than that of men, especially menopausal women. In parents with rheumatoid, the incidence rate of rheumatoid children is 2-10 times higher than that of the general population. Among close relatives with rheumatoid art...
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Patent Timeline
Login to View More IPC IPC(8): C12Q1/6883C12Q1/6869
CPCC12Q1/6883C12Q1/6869C12Q2600/156C12Q2600/16C12Q2531/113C12Q2537/143C12Q2521/319C12Q2521/525C12Q2533/101C12Q2563/107
Inventor 廖华黄宇东方浜杏
Owner 广东瑞昊生物技术有限公司