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A gene detection method and application of nucleic acid mass spectrometry-based drug for rheumatic immune diseases

A technology for rheumatic immunity and genetic testing, which is applied in biochemical equipment and methods, microbiological determination/inspection, DNA/RNA fragments, etc., can solve the problem of inability to accurately and comprehensively evaluate the effectiveness and safety of treatment, different detection methods, difficult To meet the actual clinical needs and other issues, to achieve the effect of high commercial return, low cost and short cycle

Active Publication Date: 2021-09-21
NANJING SIMCERE MEDICAL LAB CO LTD +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Although there are tests related to individualized medication for rheumatic immune diseases in the prior art, different testing methods have different emphasis, which is difficult to meet the actual clinical needs
Currently on the market, tests related to individualized medicine for rheumatic immune diseases (especially rheumatoid immune diseases) cannot accurately and comprehensively evaluate the effectiveness and safety of treatment

Method used

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  • A gene detection method and application of nucleic acid mass spectrometry-based drug for rheumatic immune diseases
  • A gene detection method and application of nucleic acid mass spectrometry-based drug for rheumatic immune diseases
  • A gene detection method and application of nucleic acid mass spectrometry-based drug for rheumatic immune diseases

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0067] Example 1 Demonstration and Screening of Detection Sites

[0068] Considering the genotype characteristics of the Chinese population, the corresponding polymorphic sites for rheumatic immune diseases (especially rheumatoid) drugs were selected, and the feasible corresponding SNP sites were established as follows: rs10306114, rs1042597, rs1045642, rs1050828, rs1050829, rs10509681, rs1051266 、rs1057910、rs1061631、rs1065852、rs10919563、rs11188072、rs1128503、rs1142345、rs11572080、rs11678615、rs116855232、rs11706052、rs12041331、rs12208357、rs12248560、rs1229984、rs1495741、rs1544105、rs16947、rs1695、rs17602729、rs1799724、rs1799971、rs1800462、rs1800629、rs1801131 、rs1801133、rs1801279、rs1801280、rs2032582、rs2187668、rs2231142、rs2234693、rs2740574、rs289714、rs3131003、rs3213422、rs3397、rs34650714、rs35167514、rs361525、rs3731722、rs3758149、rs3794271、rs41303343、rs4244285、rs4646437、rs4673993、rs4802101、rs4846051、rs4986893 、rs5030865、rs5918、rs6028945、rs6138150、rs662、rs671、rs7046653、rs719235、rs7254579、rs7300...

Embodiment 2

[0069] The design of embodiment 2 primers and the establishment of reaction system

[0070] Given that MassARRAY detection is a reaction based on multiplex PCR amplification, primer combinations must avoid issues such as cross-amplification, biased amplification, and non-specificity (D. van den Boom et al. / International Journal of Mass Spectrometry, 238(2004) , 173–188), so the design of primers for this reaction system needs a lot of optimization.

[0071] First, the present invention adjusts relevant parameters through the primer design software (Assay Design Suite) of the MassARRAY website, completes the preliminary design of primers for PCR and UEP at 77 sites, derives the designed primers and each parameter file, and synthesizes the primers. Prepare amplification primer MIX and extension primer MIX according to the primer configuration table, and fine-tune the extension primer MIX until it meets the requirements. Primer testing and optimization were then performed. Spec...

Embodiment 3

[0123] Example 3 Clinical outcome experiment (compared with generation sequencing)

[0124] After confirming the optimal reaction system, the present invention carries out a series of verification experiments, including the experimental verification of accuracy, repeatability and success rate of one experiment. The specific verification scheme is as follows:

[0125] (1) Accuracy experimental verification plan: at least 5 samples from each of the 77 sites were selected for Sanger sequencing, and the results of Sanger sequencing and MassARRAY were compared.

[0126] If the consistency is required to be greater than 95%, the verification is passed.

[0127] (2) Reproducibility experiment verification scheme: 5 clinical samples were selected, and each sample was repeated twice for detection.

[0128] The consistency of the two test results is required to be 100%.

[0129] The specific verification process is as follows: firstly, the amplification primer MIX and the extension p...

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Abstract

The invention provides a gene detection method and application of rheumatoid immune disease (especially rheumatoid disease) medicine based on nucleic acid mass spectrometry. Based on the MassArray nucleic acid mass spectrometry platform, the present invention designs and optimizes the primer set and its detection system for detecting SNP sites related to rheumatism immunotherapy, and can accurately type point mutations related to rheumatism medication, with strong accuracy, high sensitivity, good repeatability, Low cost, short detection cycle, intuitive results and many other advantages.

Description

technical field [0001] The invention relates to the field of gene detection, in particular to a nucleic acid mass spectrometry-based gene detection method for rheumatism drugs (especially rheumatoid) and its application. Background technique [0002] The most common clinical rheumatic immune diseases mainly include rheumatoid arthritis, systemic lupus erythematosus, ankylosing spondylitis, and gout. Among them, rheumatoid arthritis (RA) is a chronic systemic disease with unknown etiology, mainly inflammatory synovitis. It is characterized by polyarticular, symmetrical, and aggressive arthritis of the small joints of the hands and feet, often accompanied by involvement of extra-articular organs and positive serum rheumatoid factor, which can lead to joint deformities and loss of function. Rheumatic immune diseases are protracted and recurring diseases. Most patients need to live with the disease for a long time, and most of the diseases currently have no radical cure. The pu...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6886C12Q1/6858C12N15/11
CPCC12Q1/6858C12Q1/6886C12Q2600/106C12Q2600/156C12Q2565/627
Inventor 王应东王丽男胥慧刘世霆郑萍莫立乾赵进军林静丽李诗濛任用
Owner NANJING SIMCERE MEDICAL LAB CO LTD
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