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Micro-satellite instability multi-detection system and kit

An unstable, multiple detection technology, applied in the direction of microbial determination/inspection, biochemical equipment and methods, etc., can solve the problems such as the inability to exert the advantages of multiplex PCR experiments, insufficient clinical data support, and high experimental costs. Conducive to standardized operation, convenient for large-scale promotion and application, and intuitive effect of PCR result analysis

Active Publication Date: 2014-10-08
山东国九堂制药集团股份有限公司
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Problems solved by technology

However, the cost of the experiment is high due to a variety of fluorescent markers, and it can only be detected on a multi-channel fluorescent quantitative PCR instrument; the addition of sex-determining sites is also very limited in preventing experimental operators from confusing samples. Distinguishing between specimens does not help at all but adds unnecessary experimental expense
One of the main purposes of multiplex PCR experiments is due to cost considerations. Only by adding expensive fluorescent-labeled primers for triple PCR reactions does not give full play to the cost advantages of multiplex PCR experiments.
And the advantages of the 8 quasi-monomorphic single nucleotide repeat sites in predicting microsatellite instability status are not yet supported by sufficient clinical data

Method used

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  • Micro-satellite instability multi-detection system and kit
  • Micro-satellite instability multi-detection system and kit

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Embodiment 1

[0039] 1. The composition of the kit.

[0040] The microsatellite instability detection kit of the present embodiment comprises PCR reaction solution, primer mixture, hot start enzyme, positive control solution and negative control solution, PCR reaction solution, primer mixture, hot start enzyme, positive control solution and negative control solution The control solutions were placed in different test tubes, as shown in Table 1.

[0041] Table 1 Kit composition table

[0042]

[0043] The PCR reaction solution is prepared from 10×PCR buffer, dNTPs and hot-start enzyme. 10×PCR buffer includes 100mM Tris-HCl, 500mM KCl and 15mM MgCl 2 , the pH value of the Tris-HCl buffer used to configure the PCR buffer is 8.3. dNTPs include dATP, dGTP, dCTP and dTTP, the concentration of dATP, dGTP, dCTP and dTTP is 2.5mM, and the final concentration in the reaction system is 0.2mM. The hot start enzyme uses Taq DNA polymerase at a concentration of 5U / μl, and the final concentration i...

Embodiment 2

[0086] The rest of the microsatellite instability detection kit of this embodiment is the same as that of Example 1, except that the PCR reaction solution and the primer mixture are mixed together in advance and placed in the same test tube.

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Abstract

The invention relates to a micro-satellite instability multi-detection system and a kit. The detection system comprises a universal-specific gomphosis primer pair and a mixture of universal primers, wherein the universal-specific gomphosis primer pair respectively aims at mononucleotide micro-satellite loci BAT-26, BAT-25, NR22, NR24 and NR21; the universal primer is Flu-UP; a sequence of Flu-UP sequence is arranged at each upstream primer 5' end of the universal-specific gomphosis primer pair; and the universal primer Flu-UP is a fluorescence labeling primer. By adopting the micro-satellite instability multi-detection system and the kit provided by the invention, the defect that different primers need multiple fluorescence labels is overcome, 5 micro-satellite gene loci can be detected in one same PCR (Polymerase Chain Reaction) reaction system, the result is visible, reliable, time-saving and efficient, the cost is saved, and the MSI (Micro-satellite Instability) gene loci can be effectively detected and analyzed.

Description

technical field [0001] The invention relates to a multiple detection system and detection products thereof, in particular to a multiple detection system for microsatellite instability and detection products using the system, belonging to the field of biotechnology. Background technique [0002] Microsatellite Instability (MSI) refers to the loss of mismatch repair genes caused by DNA methylation or gene mutation, which leads to changes in the length of microsatellite repeat sequences, manifested as the same microsatellite locus between different individuals and the same individual The number of repeating units of microsatellite loci is different between normal tissues and certain abnormal tissues, so that they cannot normally play a regulatory role, resulting in abnormal cell proliferation and differentiation, and promoting the formation of malignant tumors. A large number of studies have shown that MSI is closely related to the occurrence of colorectal cancer, gastric cance...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6858C12Q2537/143C12Q2563/107C12Q2531/113
Inventor 赵新泰王明李璐
Owner 山东国九堂制药集团股份有限公司
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