Gene detection kit for detecting individual drug use of tacrolimus drug
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A gene detection and tacrolimus technology, which is applied in the field of gene detection kits for the detection of individualized drug use of tacrolimus drugs, to achieve the effects of reducing drug side effects, realizing individualized treatment, and improving drug efficacy
Inactive Publication Date: 2019-10-22
上海佰臻生物科技有限公司
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[0004] The existing technology mainly has the following technical problems: lack of a tacrolimus individualized drug gene detection kit for the Chinese population with high detection sensitivity and high accuracy
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Embodiment 1
[0025] A gene detection kit for detecting individualized drug use of tacrolimus, comprising the following components:
[0026] (1) CYP3A5*3 (-253-1A>G) and POR (1508C>T) site amplification and sequencing primers 2 pairs, each primer 1OD, primer sequence see Table 1;
[0031] This kit is stored at -20°C, and repeated freezing and thawing should be avoided as much as possible.
Embodiment 2
[0033] A method for using a gene detection kit for detecting tacrolimus drug individualized drug use, comprising the following steps:
[0034] (1) extract the genomic DNA of the sample;
[0035] (2) PCR amplification of CYP3A5*3 (-253-1A>G) and POR (1508C>T) loci
[0036] For CYP3A5*3 (-253-1A>G) and POR (1508C>T) sites, design primers according to Table 1 for PCR amplification. Each reaction system has a total volume of 16 μL, including 7.5 μL of Taq enzyme mixture (dNTP, 10×PCR reaction buffer, MgCl2), 5.5 μL of deionized water, primer pair (10uM) 1 μL, genomic DNA (100ng / ul) 1ul ;The reaction condition is 95°C for 15min, 14 cycles (-0.5°C / cycle) of 94°C for 30sec, 63°C for 30sec, 72°C for 45sec; then 30 cycles of 94°C for 30sec, 56°C for 30sec, 72°C for 45sec, Finally, 72°C for 10min;
[0037] (3) Purification of PCR products
[0038] Each reaction has a total volume of 5.6 μL. 4 μL of PCR product was added, and then 1.6 μL of SAP enzyme mixture (SAP enzyme, ExoI enzym...
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Abstract
The invention provides a gene detection kit for detecting individualized drug use of a tacrolimus drug. The kit comprises the following components including two pairs of PCR amplification and sequencing primers for CYP3A5*3 (-253-1A>G) and POR (1508C>T) site regions, a PCR amplification reagent, a PCR product purification reagent and a DNA sequencing reagent. According to the sequencing primers for the CYP3A5*3 (-253-1A>G) site, the forward primer is TGCCCTTGCAGCATTTAG, and the reverse primer is ACACCCAGGAAGCCAGAC; according to the sequencing primers for the POR (1508C>T) site, the forward primer is GGTGGTTGTGGAGTACGAGA, and the reverse primer is CGCTCCTGGATGAAGCCTAT. The kit is used for genotyping detection of the CYP3A5*3 (-253-1A>G) and POR (1508C>T). By applying the kit in detection, the individual drug metabolism type including a normal metabolite and a slow metabolism type can be identified, the drug dosage is adjusted, the drug efficacy is improved, the toxic and side effects ofthe drug are reduced, and individualized treatment is realized.
Description
technical field [0001] The invention relates to the field of biotechnology, in particular to a gene detection kit for detecting individualized drug use of tacrolimus. Background technique [0002] The calcineurin inhibitor tacrolimus is a macrolide antibiotic isolated from the metabolites of Streptomyces and was used clinically in 1989. It mainly inhibits the activation process of T cells by specifically inhibiting the activity of neural calcineurin. At the same time, it also produces T cell-dependent antibodies that cyclosporine does not have, and blocks the transcription of cytokines, presenting a double effect. Immunosuppressive effect. Tacrolimus is mainly used for graft rejection after liver or kidney transplantation. However, due to the narrow therapeutic window of tacrolimus and the large individual differences in pharmacokinetics, it is administered according to the conventional dose, and some patients may suffer from "insufficient immunosuppression" due to too low...
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