Single nucleotide polymorphism of human WNK4 genes and application thereof
A single nucleotide polymorphism and gene technology, applied in the fields of molecular biology and medicine, can solve the problem of unproven rs2290041 locus correlation
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Embodiment 1
[0036] Fluorescent PCR detection
[0037] 1. Experimental materials
[0038] The 7900HT fluorescence quantitative PCR instrument was purchased from ABI Company of the United States, and the polymerase chain reaction solution (TaqMan EXPressMaster Mix) was custom-synthesized by Applied Biosystems (ABI).
[0039] 2. Primer and probe design and synthesis:
[0040] Using the partial sequence of exon 14 of the WNK4 gene as a template, TaqMan primers and probe sites were analyzed using Primer ExpressTM 2.0 software, and custom-synthesized by Applied Biosystems (ABI).
[0041] Primers for detection:
[0042] WNK4 gene rs2290041 upstream primer sequence: 5′-GAGTGCAGAGTGGAGTCTCCCCTAATC-3′
[0043] WNK4 gene rs2290041 downstream primer sequence: 5′-CTCACTGGCTGTGATGACTGTGG-3′
[0044] Fluorescent probes:
[0045] WNK4 gene rs2290041 fluorescent probe 1: 5′-VIC-AGGCTAGAGAGGGGACTA-TAMRA-3′
[0046] WNK4 gene rs2290041 fluorescent probe 2: 5′-FAM-AGGCTAGGGAGGGGACTA-TAMRA-3′
[0047] ...
Embodiment 2
[0059] The rs2290041 site of the WNK4 gene, a susceptibility gene of essential hypertension, was detected by sequencing. 10 cases of each of the above-mentioned hypertension cases and control groups were selected for sequencing to determine the genotype of rs2290041.
[0060] 1. Experimental method
[0061] The above-mentioned fluorescent PCR primers were still used as primers for PCR sequencing, and the amplified products were directly sequenced after purification. The sequencing instrument is ABI's 3130xl genetic analyzer, which is analyzed with sequence analysis5.2 analysis software, and the results can also be viewed with chromas.
[0062] 2. The experimental results showed that the sequencing results of 20 cases were completely consistent with the genotype analysis results of 7900 fluorescent PCR.
[0063] 3. Association analysis of WNK4 gene rs2290041 genotype and hypertension susceptibility
[0064] Comparison of distribution of WNK4 gene rs2290041 in hypertensive pa...
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