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SNP rs4646418 of CYP1A2 gene and application thereof in relevant drug metabolism activity detection

A CYP1A2, P4501A2 technology, applied in the field of molecular biology and medicine, can solve the problem of unclear drug metabolism activity or ability related, not affecting CYP1A2 polymorphism and other problems

Inactive Publication Date: 2010-06-23
CHINESE NAT HUMAN GENOME CENT AT SHANGHAI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0016] In summary, although it is known in the art that polymorphisms of CYP1A2 enzymes or CYP1A2 genes are associated with the ability to metabolize certain drugs, people have not yet affected those specific polymorphisms (including SNPs) in CYP1A2, and it is not clear Which polymorphisms (including SNPs) are associated with drug metabolism activity or capacity

Method used

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  • SNP rs4646418 of CYP1A2 gene and application thereof in relevant drug metabolism activity detection
  • SNP rs4646418 of CYP1A2 gene and application thereof in relevant drug metabolism activity detection
  • SNP rs4646418 of CYP1A2 gene and application thereof in relevant drug metabolism activity detection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0069] Experimental materials and methods

[0070] 96 normal Chinese liver samples were collected clinically. The samples were collected from the pathological biopsy part of normal liver donors, about 50-200 mg per case, immediately immersed in 1 mL RNALater after collection, and kept overnight at 4°C. After the samples were fully soaked by RNALater, they were transferred to -70°C for storage. Ethics approval was obtained for the collection and use of working samples for this study.

[0071] Samples stored at -70°C were reconstituted at room temperature, removed from the RNALater, and placed in DEPC-treated ddH 2 RNALater was rinsed in O, transferred to 1 mL Trizol (Invitrogen Inc.), and homogenized with Pro200homogenizer (Pro scientific Inc.). DNA and RNA were then obtained according to Trizol's protocol. Preliminary quantification was performed with a Biophotometer (Eppendorf Inc.). The obtained RNA was treated with DNase I (Takara Inc.), purified by extraction with phen...

Embodiment 2

[0085] Validation of SNP rs4646418

[0086] 2.1 DNA extraction

[0087] DNA was extracted from human blood by the conventional phenol-chloroform method, and the concentration was corrected to 20ng / ul for conventional PCR amplification.

[0088] 2.2 Design of PCR and sequencing primers

[0089] According to the sequence of CYP1A2 gene shown in SEQ ID NO:1, the following primers were designed and synthesized. The specific primers are shown in Table 1 below.

[0090] Table 1 Primer sequence list

[0091] Primer name

Sequence (5'-3')

SEQ ID NO:

sense primer

gggattggag agaaaggtgg cggag

2

antisense primer

ctctctccgc aaggccttcc ctgac

3

[0092] 1.2.3 PCR amplification of CYP1A2 gene

[0093] Using the extracted DNA as a template, PCR amplification was performed on a GeneAmp 9700 PCR instrument with the Touchdown program using Taq enzyme. The reaction conditions are: pre-denaturation at 94°C for 2 minutes, denaturation at 94°...

Embodiment 3

[0099] Individual drug metabolism activity assays

[0100] Because it is known that about 90% of caffeine is metabolized by CYP1A2 in the human body, there is a correlation between the activity of CYP1A2 and the metabolism of caffeine.

[0101] In this example, the method of Example 2 was used to genotype the CYP1A2 gene based on the SNP site at position 830 T→C in SEQ ID NO: 1 in healthy Chinese population. And observe the relationship between different genotypes and the ratio of plasma caffeine metabolism.

[0102] Select 20 healthy subjects (10 males and 10 females) aged 18-30 years old (mean age 21 ± 2 years old) to participate in this test, wherein in SEQ ID NO: 1: 830 are T and C 10 each. The entire experimental process was carried out in accordance with the National Human Genome Research Ethics Guidelines, and informed consent was obtained from all subjects.

[0103] All subjects were non-smoking individuals, and had not consumed coffee, tea, Coca-Cola, chocolate or ...

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Abstract

The invention discloses SNP rs4646418 of a CYP1A2 gene and application thereof in relevant drug metabolism activity detection, and also provides a method for prejudging drug metabolism activity. The method comprises the steps of detecting whether the cytochrome oxidase P4501A2 gene (CYP1A2) and a transcript of a human individual have variation or not compared with a normal cytochrome oxidase P4501A2 gene and a normal transcript, and if so, showing that the drug metabolism activity of the individual is different from common or normal people, for the individual. The invention also discloses a corresponding detection kit used for prejudging the drug metabolism activity of the individual.

Description

technical field [0001] The present invention relates to the fields of molecular biology and medicine. More specifically, it relates to the single nucleotide polymorphism (single nucleotide polymorphism, SNP) of cytochrome oxidase P4501A2 gene (Cytochrome P4501A2, CYP1A2) and its correlation with the detection of drug metabolism activity. The invention also relates to methods and kits for detecting said SNP. Background technique [0002] Individual differences in drug effects are already a common phenomenon, and the drug ineffectiveness or severe drug side effects caused by this difference has attracted more and more attention from clinicians and pharmacogeneticists. Differences in drug metabolism ability and drug effect are one of the main reasons for individual differences in drug effect. Some drug metabolism and effect-related genes have been found to have not only DNA sequence variation in the population, but also large individual differences in their RNA expression leve...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/53
Inventor 黄薇王大志王蓓兰王志敏雷蓉施锦绣
Owner CHINESE NAT HUMAN GENOME CENT AT SHANGHAI
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