Method for detecting transcription level of Macaca mulatta gene SLC22A11/OAT4 through RT-qPCR

A technology of gene transcription and gene expression level, applied in the field of detection, to achieve the effect of high specificity, strong specificity and high sensitivity

Pending Publication Date: 2018-07-27
INST OF MEDICAL BIOLOGY CHINESE ACAD OF MEDICAL SCI
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  • Abstract
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Problems solved by technology

At present, no member of the organic anion transporter family has been used in rhesus monkeys at home and abroad. SLC22A11 / Study on Quantitative Detection Method of OAT4 Transcription Level

Method used

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  • Method for detecting transcription level of Macaca mulatta gene SLC22A11/OAT4 through RT-qPCR
  • Method for detecting transcription level of Macaca mulatta gene SLC22A11/OAT4 through RT-qPCR
  • Method for detecting transcription level of Macaca mulatta gene SLC22A11/OAT4 through RT-qPCR

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Embodiment

[0052] RT-qPCR detection in macaques SLC22A11 / The method for OAT4 gene transcription level comprises the following steps:

[0053] (1) Design the following primers:

[0054] According to the macaca mulatta monkey in the NCBI gene bank SLC22A11 / OAT4 Nucleotide sequence number: NM_018484, macaque ( macaca mulatta )of GAPDH Nucleotide sequence number NM_001195426.1 designed primers, the above primers were synthesized by Shanghai Bioengineering Co., Ltd.:

[0055] Macaque SLC22A11 / The specific upstream and downstream primers for OAT4 gene expression level are:

[0056] SLC22A11 / OAT4 F: 5'-GGTGGCTGATTATTAAGGGCAAAC-3'

[0057] SLC22A11 / OAT4 R: 5'-CTTCACGCTGGACATCAGCA-3'

[0058] The specific upstream and downstream primers of the rhesus monkey GAPDH gene as an internal reference gene are:

[0059] GAPDH F: 5'-AGCCCCATCACCATCTTCC-3'

[0060] GAPDH R: 5'-AATGAGCCCCAGCCTTCTC-3';

[0061] Macaque SLC22A11 / OAT4 The primer sequences and fragment sizes for quan...

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Abstract

The present invention provides a method for detecting the transcription level of a Macaca mulatta gene SLC22A11 / OAT4 through RT-qPCR. According to the method, cDNA obtained through the reverse transcription of the total RNA of a sample is used as a template, qPCR amplification is performed with a PCT primer combination to obtain the amplification product of the Macaca mulatta gene SLC22A11 / OAT4 fragment, the normalized gene folds are expressed by using zero point as a control to obtain a relative expression value, the relative expression values of the mRNA of the Macaca mulatta gene SLC22A11 / OAT4 under different physiological conditions can be calculated according to the relative expression value, and the transcription level change of the Macaca mulatta gene SLC22A11 / OAT4 can be quantitatively detected according to the relative expression values so as to provide the effective way for the research on the function of Macaca mulatta gene SLC22A11 / OAT4 and the influence caused by related drugs. According to the present invention, the method is suitable for the real-time quantitative PCR detection, and has advantages of simple operation, high reproducibility, low detection cost, high sensitivity and strong specificity.

Description

technical field [0001] The present invention relates to a detection method, in particular to a real-time fluorescent quantitative RT-qPCR method for detection of macaque organic anion transporter family members SLC22A11 / The method for the transcription level of OAT4 gene belongs 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. in SLC2A9 , SLC22A11 and SLC22A12 Loss-of-function mutations in genes cause hereditary hypouricemia, while overexpression enhances uric acid reabsorption. ABCG2, SLC17A1 and SLC17A3 Defective gene function variants reduce renal and intestinal excretion of uric acid. Members of the organic anion transporter family SLC22A11 / OAT4 is expressed in proximal tubule cells and anchored to the apical membrane. OAT4 has been shown to be an asymmetric low-affin...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6851C12Q1/6888C12Q2600/158C12Q2600/166C12Q2531/113C12Q2545/101C12Q2563/107
Inventor 唐东红王陈芸叶尤松李哲丽肖涵邱炳玲陈倩杨浩杨忠马开利鲁帅尧
Owner INST OF MEDICAL BIOLOGY CHINESE ACAD OF MEDICAL SCI
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