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RT-qPCR detection method of slc22a12/urat1 gene transcription level in macaques

A gene transcription, macaque technology, applied in the field of detection, to achieve the effect of high repeatability, simple method and high sensitivity

Active Publication Date: 2021-05-14
INST OF MEDICAL BIOLOGY CHINESE ACAD OF MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, there is no research on the quantitative detection method of SLC22A12 / URAT1 gene transcription level using rhesus monkey at home and abroad

Method used

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  • RT-qPCR detection method of slc22a12/urat1 gene transcription level in macaques
  • RT-qPCR detection method of slc22a12/urat1 gene transcription level in macaques
  • RT-qPCR detection method of slc22a12/urat1 gene transcription level in macaques

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Embodiment

[0056] RT-qPCR detection in macaques SLC22A12 The method for the transcription level of / URAT1 gene is characterized in that it comprises the following steps:

[0057] (1) Design the following primers:

[0058] According to the cynomolgus monkey in the NCBI gene bank ( Macaca fascicularis )of SLC22A12 / URAT1, The gene accession number is AB738914.1, primers were designed with Primer Premier 5.0 software, synthesized by Bao Biology Co., Ltd. GAPDH As an internal reference gene, macaque ( Macaca mulatt )of GAPDH Nucleotide sequence design primers, gene accession number NM_001195426.1.

[0059] The specific upstream and downstream primers for the expression level of macaque SLC22A12 / URAT1 gene are:

[0060] SLC22A12 / URAT1 F:5'-TCAGCCATGAGGGAGGAAC-3'

[0061] SLC22A12 / URAT1 R:5'-CCAAAGGCGAACCAGCA-3';

[0062] The specific upstream and downstream primers of the rhesus monkey GAPDH gene as an internal reference gene are:

[0063] GAPDH F: 5'- AGCCCCATCACCATCTT...

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Abstract

The present invention provides a kind of RT-qPCR detection rhesus monkey SLC22A12 / URAT1 gene transcription level method. The cDNA of rhesus monkey kidney tissue obtained by reverse transcription synthesis of total RNA extracted from fresh kidney tissue of rhesus monkey was used as a template, and real-time fluorescence quantitative PCR was amplified by PCR primer combination to obtain SLC22A12 / URAT1 gene fragment and reference gene GAPDH The Ct value, dissolution peak and amplification efficiency of the fragment; after processing, the normalized SLC22A12 The ΔΔC(t) value of the multiple expression of the / URAT1 gene was obtained SLC22A12 / Relative expression value of URAT1 gene transcript level. It provides an effective way to study the function of macaque urate anion transporter 1 and the influence of related drugs on it. The invention is suitable for real-time quantitative PCR detection, and has high sensitivity, strong specificity, simple operation and high repeatability.

Description

technical field [0001] The invention relates to a detection method, in particular to a method for detecting the gene transcription level of macaque urate anion transporter 1 (SLC22A11 / URAT1) by real-time fluorescence quantitative method (RT-qPCR), belonging to the field of molecular biology technology. Background technique [0002] In recent years, genome-wide association studies (GWAS) have detected multiple susceptibility loci and related candidate genes that lead to hyperuricemia and gout. Loss-of-function mutations in SLC2A9, SLC22A11, and SLC22A12 genes can cause hereditary hypouricemia, while overexpression can enhance uric acid reabsorption. Defective-function variants in the ABCG2, SLC17A1, and SLC17A3 genes reduce renal and intestinal excretion of uric acid. Urate anion transporter 1 (urate anion transporter 1, URAT1) is the main transporter for human renal tubular reabsorption of uric acid, and is the first protein found to be related to renal urate transport, mai...

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6851
Inventor 唐东红叶尤松王陈芸李哲丽邱炳玲肖涵陈倩杨浩杨忠马开利鲁帅尧
Owner INST OF MEDICAL BIOLOGY CHINESE ACAD OF MEDICAL SCI