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Kit for detecting thiopurine methyltransferase genotype, method and application thereof
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A mercaptopurine methyl and kit technology, applied in the biological field, can solve the problems of treatment failure, treatment interruption, and high disease recurrence rate, and achieve the effects of improving amplification efficiency and saving time.
Inactive Publication Date: 2009-12-02
BEIJING HUAANFO BIOMEDICAL RES CENT +1
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Individuals with high TPMT activity have a low concentration of active metabolites-mercaptoguanine phosphates (TGNs) after medication, so the recurrence rate of the disease is high; and for individuals with TPMT deficiency, medical conventional doses of purine drugs can also cause in vivo The accumulation of mercaptoguanine phosphate produces serious side effects such as severe bone marrow suppression and liver damage, which eventually leads to the forced interruption of treatment, resulting in treatment failure (Coulthard SA, Howell C, Robson J, et al.Blood, 1998 , 92: 2856-2862; Relling M V, Hancock ML, Boyett JM, et al. Blood, 1999, 93: 2817-2823)
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Embodiment 1
[0076] The present invention will be further described in detail below by taking the TPMT G460A wild-type positive controlplasmid as an example.
[0077] 1. Primers and templates
[0078] Gene fragment amplification uses the following primers:
[0079] TPMT 460 For: 5'-CAGCGTAGTTGGTGTGGAAA-3';
[0080] TPMT 460 Back: 5'-GCGATCACCTGGATTAATGG-3'.
[0081] The template uses human genomic DNA (No. AI011611, concentration 15ng / μl)
[0093] 94°C for 3 minutes; 94°C for 30 seconds, 63°C for 45 seconds, 72°C for 30 seconds, 35 cycles; 72°C for 7 minutes.
[00...
Embodiment 2
[0125] The present invention will be further described in detail below by taking the TPMT genotype detection process of leukemia patient genes as an example.
[0126] We used this kit to detect the mercaptopurine methyltransferase genotypes of 20 patients, the process is as follows:
[0128] Since the sample is genomic DNA, use a UV spectrophotometer to detect the DNA concentration of the sample, and dilute the DNA concentration to 30-100ng / ml
[0129] 2. Configuration of G460A-specific PCR amplification mixture
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technical field [0001] The invention belongs to the field of biotechnology, and relates to a mercaptopurine methyltransferase genotype detection kit, method and application. More specifically, the present invention relates to a kit, method and application for analyzing the polymorphism of a gene locus encoding mercaptopurine methyltransferase by polymerasechain reaction method. Background technique [0002] Purine drugs, including 6-mercaptopurine (6-MP), 6-mercaptopurine (6-TG) and mercaptopurine (AZA) are commonly used clinical tumor chemotherapy drugs. They interfere with the metabolism of nucleic acids in the body, including Both de novo and salvage synthesis pathways of nucleic acids come into play. Purine drugs are drugs without intrinsic biological activity, and must undergo a series of metabolism in the body before they can exert their anti-leukemic effect. [0003] Studies have shown that the efficacy of purine drugs is closely related to the activity of thiopuri...
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