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Application of abcc3-013 mRNA in the preparation of a kit for detecting clopidogrel resistance and its kit

一种氯吡格雷抵抗、试剂盒的技术，应用在生物化学设备和方法、微生物的测定/检验、含有效成分的医用配制品等方向，能够解决缺少氯吡格雷抵抗标记物、氯吡格雷抵抗机制不十分清楚等问题

Active Publication Date: 2020-10-09
PULING BIOLOGY (NANJING) CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The mechanism of clopidogrel resistance is not very clear, and clinically effective markers of clopidogrel resistance are also lacking

Method used

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  • Application of abcc3-013 mRNA in the preparation of a kit for detecting clopidogrel resistance and its kit
  • Application of abcc3-013 mRNA in the preparation of a kit for detecting clopidogrel resistance and its kit
  • Application of abcc3-013 mRNA in the preparation of a kit for detecting clopidogrel resistance and its kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Example 1: Application of ABCC3-013 mRNA in the preparation of a kit for detecting clopidogrel resistance.

[0021] 1. A kit for detecting clopidogrel resistance, including the following components:

[0022] TRIZOL reagent,

[0023] Chloroform,

[0024] isopropanol,

[0025] DEPC water,

[0026] Oligo dT primer (Oligo dT primer),

[0027] Random 6-nucleotide primers (Random 6-mers),

[0028] reverse transcriptase (PrimeScript TM RT Enzyme Mix),

[0029] RNase-free water (RNase-free dH 2 O),

[0030] Reverse transcription buffer (PrimeScript TM buffer),

[0031] Taq enzyme, dNTPs, Mg 2+ , SYBR Green I (SYBR Premix Ex Taq TM ),

[0032] ROX Reference Dye, and

[0033] Specific primers SEQ ID NO.5 and SEQ ID NO.6 for ABCC3-013 mRNA (nucleotide sequence shown in SEQ ID NO.2).

[0034] 2. How to use the kit

[0035] The peripheral blood RNA of the patient was extracted, and the content of ABCC3-013 mRNA was detected by qRT-PCR, and the specific operation wa...

Embodiment 2

[0062] Example 2: Experimental research.

[0063] ABCC3-013 mRNA is a non-wild-type mRNA splicing variant of ABCC3 gene due to abnormal mRNA splicing caused by point mutations. In the following experiments, ABCC3-002 mRNA, another splice variant of the ABCC3 gene, was used as a control to examine the biological activity of ABCC3-013 mRNA.

[0064] 1. Cell culture: HepG2 cells were cultured in high-glucose DMEM medium containing 10% inactivated fetal bovine serum, 50 U / ml penicillin and 50 μg / ml streptomycin, at 37°C and 5% CO in saturated humidity 2 Incubator cultivation.

[0065] 2. Plasmid construction.

[0066] Method: ABCC3-002 (SEQ ID NO.1) and ABCC3-013 (SEQ ID NO.2) fragments were chemically synthesized, fragments ABCC3-002-EGFP and ABCC3-013-EGFP containing EGFP were synthesized, and random fragments were synthesized as a control group, the synthetic fragments were cloned into the TV5 vector. The recombinant vectors were named TV5, ABCC3-002, ABCC3-013, TV5-EGFP, A...

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Abstract

The present invention provides an application of ABCC3-013 mRNA in preparing a kit for detecting clopidogrel resistance. The nucleotide sequence of ABCC3-013 mRNA is shown in SEQ ID NO.2. It can detect the clopidogrel resistance of patients by detecting the expression level of ABCC3-013 mRNA in patients (such as peripheral blood), and provide a new method for assisting clinical treatment, judging prognosis and curative effect. The present invention also provides a kit for detecting clopidogrel resistance, which can detect the clopidogrel resistance of a patient by detecting the expression level of ABCC3-013 mRNA in the patient's body (for example, peripheral blood). Clinical trials have shown that its diagnostic sensitivity is 52%, and its specificity is 86%.

Description

technical field [0001] The invention relates to a kit for detecting drug resistance, in particular to a kit for detecting clopidogrel resistance. Background technique [0002] As an antiplatelet drug, clopidogrel is the most commonly used drug to prevent stent thrombosis after percutaneous coronary intervention (PCI) implantation of stents in patients with coronary heart disease. It is combined with aspirin to form "dual antiplatelet therapy" The regimen is the "gold standard" antiplatelet therapy regimen for acute coronary syndrome and PCI after stent implantation. Clopidogrel resistance (or tolerance) occurs in 10-40% of patients treated with clopidogrel, showing that the antiplatelet effect of clopidogrel does not meet the expected requirements, resulting in poor efficacy of the drug. [0003] The existing platelet aggregation test and thromboelastography can evaluate clopidogrel resistance to a certain extent; however, the repeatability of the platelet aggregation test ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6883
CPCA61K31/7105C12Q1/6883C12Q2600/106C12Q1/68C12Q1/6816C12Q1/6827C12Q1/6832C12Q1/6876
Inventor 谢红光柳伟强潘玉琴杨平殷蒨米琼宇
Owner PULING BIOLOGY (NANJING) CO LTD
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