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Primers, probes and fluorescent pcr kits for detecting ugt1a1 genotype and gst gene deletion type of neonatal jaundice

A UGT1A1, kit technology, applied in biochemical equipment and methods, DNA/RNA fragments, recombinant DNA technology, etc., can solve the problems of low detection throughput, long detection period, inapplicability of multiple genes and multiple sites, etc. Intuitive and accurate interpretation, short detection time, the effect of amplification system and program optimization

Active Publication Date: 2021-11-30
昆明和合医学检验所有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] At present, the main genes for the detection of neonatal jaundice are UGT1A1*6 and UGT1A1*28. The detection methods mainly include serum bilirubin measurement, serum PCT detection and PCR-Sanger sequencing. Although the traditional PCR-Sanger sequencing method is Recognized as the "gold standard" for genotyping detection, but the detection cycle is long and the detection throughput is low, so it is not suitable for the detection of multiple genes and multiple loci. Compared with ordinary PCR, real-time fluorescence quantitative polynucleotide Chain reaction (Real-time Quantitative polymerase chain reaction, RT-PCR) draws a dynamic change graph for the cumulative rate of amplified DNA during the entire PCR process, eliminating the problem of a large coefficient of variation when determining the abundance of end products, and its It has the advantages of accurate typing, simple and fast operation, good accuracy, high sensitivity, support for batch detection and wide application, etc.

Method used

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  • Primers, probes and fluorescent pcr kits for detecting ugt1a1 genotype and gst gene deletion type of neonatal jaundice
  • Primers, probes and fluorescent pcr kits for detecting ugt1a1 genotype and gst gene deletion type of neonatal jaundice
  • Primers, probes and fluorescent pcr kits for detecting ugt1a1 genotype and gst gene deletion type of neonatal jaundice

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] Example 1 Polymorphism detection of neonatal jaundice-related genes

[0055] (1) Obtain the genomic DNA of the sample to be tested:

[0056]Collect fresh peripheral whole blood samples from neonatal jaundice of the same age group, and use TianGen Blood / Cell / Tissue Genomic DNA Extraction Kit (DP304) to extract genomic DNA, and use NP80-touch (Germany IMPLEN) to measure the concentration and purity of DNA. Genomic DNA is then preserved.

[0057] (2) Design of specific primers and probes:

[0058] According to literature research, two polymorphic sites of 6 genes of UGT1A1 gene and GST gene related to neonatal jaundice were selected, as shown in Table 1 and Table 2 above, and the DNA sequences of each polymorphic site were searched through NCBI, and designed and To synthesize the amplification primers of the embodiments of the present invention, for the design of primers, online / offline databases or software such as Primer Quest, Primer Premier 5.0, DNAMAN, etc. can be u...

Embodiment 2

[0120] Example 2 The assembly and use method of the polymorphism detection kit of neonatal jaundice-related genes

[0121] The kit of the present invention includes: primers and probes, internal standard system, RT-PCR reaction solution system, positive control sample, negative control, according to the usage of each component of the RT-PCR reaction solution system, calculate 24 people and 48 people The dosage of each component of the two specifications is used to prepare and assemble the components in each tube of the kit. The specific components and proportioning process of the reaction are as follows:

[0122] 1. Primers and probes: Design and synthesize 5 sets of specific primers and 2 sets of specific probes:

[0123] 5 sets of specific primers: UGT1A1 G211A upstream primer (SEQ ID NO:1), UGT1A1 G211A downstream primer (SEQ ID NO:2), UGT1A1*28 upstream primer (SEQ ID NO:3), UGT1A1*28 downstream primer (SEQ ID NO:3) ID NO:4), GSTM1 upstream primer (SEQ ID NO:5), GSTM1 do...

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Abstract

The invention provides a primer, a probe and a fluorescent PCR kit for simultaneously detecting the UGT1A1 genotype and the GST gene deletion type of genes related to neonatal jaundice. The provided primers and probes can detect UGT1A1 gene *6 type and *28 type, GST gene GSTT1 type and GSTM1 type two polymorphic sites and two deletion nucleotide types on the real-time fluorescent quantitative PCR technology platform . Using the primers, probes, and kits, the detection results have the characteristics of rapidity, high efficiency, accuracy, convenience, high sensitivity, good specificity, and low cost.

Description

technical field [0001] The invention relates to the technical field of genotype detection, in particular to a polymorphism detection kit, primers, reaction system and probes of genes related to neonatal jaundice types (physiological and pathological). Background technique [0002] Neonatal jaundice is one of the most common clinical symptoms in the neonatal period. It is a disease characterized by yellow staining of the skin, mucous membranes and sclera due to the abnormal metabolism of serum bilirubin in newborns, which causes the level of bilirubin in the blood to increase. Neonatal jaundice is divided into physiological and pathological. Physiological jaundice refers to temporary jaundice caused solely by the characteristics of bilirubin metabolism. When serum bilirubin exceeds the diagnostic criteria of physiological jaundice, it can develop into pathological hyperthyroidism. Bilirubinemia (that is, pathological jaundice), if not diagnosed and treated in time, may cause ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6883C12Q1/6858C12N15/11
CPCC12Q1/6858C12Q1/6883C12Q2600/156C12Q2600/16C12Q2600/166C12Q2531/113C12Q2561/101C12Q2563/107C12Q2537/143C12Q2545/101C12Q2545/113
Inventor 吕小波李泽东刘芳黎黄和飞谷叶李佳兴
Owner 昆明和合医学检验所有限公司
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