Primer, probe, fluorescence PCR kit and method for detecting human UGT1A1 gene polymorphism

A technology for gene polymorphism and detection primers, which is applied in the field of biomedical clinical molecular detection, can solve the problems of cumbersome data analysis, long cycle and high price

Inactive Publication Date: 2015-03-04
SUZHOU KUANGYUAN MOLECULAR BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] Aiming at the deficiencies of the prior art, such as high price, long period, low efficiency, cumbersome data analysis, etc. when detecting the UGT1A1 gene polymorphism, the present invention provides a real-time fluorescent quantitative PCR technology platform based primers and fluorescence detection method for UGT1A1 gene polymorphism detection. The probe, kit and detection method are used to detect the c.221 site (G / A) polymorphism of human UGT1A1*6 type and the *28 type promoter (TA6 / 7) polymorphism. Low, with extensive promotion value

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  • Primer, probe, fluorescence PCR kit and method for detecting human UGT1A1 gene polymorphism
  • Primer, probe, fluorescence PCR kit and method for detecting human UGT1A1 gene polymorphism
  • Primer, probe, fluorescence PCR kit and method for detecting human UGT1A1 gene polymorphism

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0109] Example 1. Design of primers and probes:

[0110] According to the UGT1A1 gene reference sequence (NG_009254) released by the National Center for Biotechnology Information NCBI's nucleic acid sequence database GeneBank and the c.221 site (G / A) polymorphism (rs4148323) published by the dbSNP database and the UGT1A1 gene *28 type TA6 / 7 polymorphism (rs34983651) information, using the Primer Express 3.0 software of ABI company to design primers and detection of UGT1A1 gene c.221 site (G / A) polymorphism and 28 type Probes, as follows:

[0111] The sequences of primers and probes for detecting the polymorphism of UGT1A1 gene *6 type c.221 site (G / A) are as follows:

[0112] Reverse primer:

[0113] UGT1A1 *6G: 5'-GTCTTCAAGGTGTAGCATGCACC-3';

[0114] UGT1A1 *6A: 5'-GTCTTCAAGGTGTAGCATGCACT-3';

[0115] Forward primer:

[0116] UGT1A1 *6F: 5’-ACTGTTGATCGCAGTGGATGG-3’

[0117] Fluorescent probes:

[0118] TM-UGT1A1*6: 5' FAM-CTAGCACCTGACGCCTCGTTGT-3' BHQ1.

[0119] The s...

Embodiment 2

[0133] Example 2. Preparation of primers

[0134] The designed primers and probe sequences are handed over to the synthesis company for synthesis, generally using automatic chemical synthesis with instruments, and a synthesis inspection report is required.

Embodiment 3

[0135] Example 3: Preparation of a kit for detecting human UGT1A1 gene polymorphisms by fluorescent PCR

[0136] Prepare two specific reaction solutions for detection of human UGT1A1 gene c.221 site (G / A) polymorphism: UGT1A1 *6G PCR reaction solution and UGT1A1 *6A PCR reaction solution; detection of human UGT1A1 gene *28 type TA6 / 7 Two specific reaction solutions for polymorphism; UGT1A1 *28TA6 PCR reaction solution and UGT1A1 *28TA7 PCR reaction solution; the PCR reaction solution contains not only buffers, magnesium ions, dNTP and other substances necessary for PCR reactions, but also detection Primers and probes and internal control primers and probes. The concentration of the PCR reaction solution components and the sequence information of the included primers and probes for the above-mentioned specific detection are as follows:

[0137] Table 1. Components of PCR reaction solution

[0138]

[0139] (1) UGT1A1 *6G PCR reaction solution

[0140] UGT1A1 *6G: 5'-GTCTTCA...

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Abstract

The invention provides a primer, a probe, a fluorescence PCR kit and a method for detecting nucleotide polymorphism at *6-type and *28-type loci of the UGT1A1 gene. According to the allele specific PCR principle, the nucleotide type at *6-type and *28-type polymorphism loci of the methylenetetrahydrofolate reductase UGT1A1 gene is detected on a real-time fluorescence quantitative PCR technical platform.

Description

technical field [0001] The invention belongs to the field of biomedical clinical molecular detection, and in particular relates to primers, probes, fluorescent PCR kits and detection methods for detecting nucleotide polymorphisms at two sites of UGT1A1 gene *6 type and *28 type. Background technique [0002] Uridine diphosphate glucuronyltransferases (UGTs) are a large class of membrane-bound enzymes that can catalyze the binding of glucuronic acid to nucleophilic substrates, and mainly exist in the endoplasmic reticulum of liver and extrahepatic tissues. UGT is divided into two families, and 17 human genes have been found to encode UGTs with different sequences. The expression of the enzyme has obvious tissue specificity, and it is mainly expressed in the liver, and other tissues include the brain, kidney, and gastrointestinal tract. UDP-glucosyltransferase 1A1 (UGT1A1 for short) is involved in the glucuronylation of various substances. The purpose of this combination reac...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6858C12Q1/6876C12Q2531/113C12Q2563/107
Inventor 王进吴梦华吴子侠何晓辉赵小龙彭飞
Owner SUZHOU KUANGYUAN MOLECULAR BIOTECH
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