Relevance Studies Of VDR Gene Promoter DNA Methylation With Idiopathic Hypocitraturia In Bai People Of Yunnan

Objective:To detect the methylation level of the VDR gene promoter in Yunnan Bai people with idiopathic hypocitrateuria,and to analyze the association between the VDR gene promoter methylation and the incidence of idiopathic hypocitrateuria in Yunnan Bai people,To explore the etiological characteristics of urinary calculi and idiopathic hypocitrateuria from the aspect of epigenetics.Methods:We selected 30 samples,of which 15 samples from the experimental group were patients with idiopathic hypocitrateuria of Yunnan Bai nationality,and the genotype of their VDR gene rs2228570(Fok Ⅰ)was FF,and 15 samples from the control group were urine citric acid.Physical examination patient of Yunnan Bai nationality with normal acid content.After the sample to be tested is treated with sulfite,the relevant primers are designed and synthesized,and then the target gene sequence is obtained through PCR amplification reaction.The T7 promoter sequence is introduced into the target sequence,and the target sequence can be obtained through the in vitro transcription process of T7 RNA polymerase.The RNA product of the sample is processed by base-specific enzyme digestion to obtain small RNA fragments that can be analyzed and detected,and the molecular weight of each fragment is detected by flight mass spectrometry.Finally,the EpiTYPER program completes the automatic processing of the data and reports the degree of methylation at each detection site.Results:on the statistical results of general data,there was no statistical difference in age,body weight,body mass index,urine volume,urine calcium content,blood PTH、blood calcium,except for urine citrate excretion(P<0.001)between 15 patients with idiopathic low citrateuria and 15 controls in the chinese bai nationality.In the statistical results of DNA methylation level,on the VDR Ⅰ fragment,the methylation level of the experimental group was 4.136%(1.655%,5.152%)higher than the methylation level of the control group 1.261%(0.827%,1.930%),and this difference is already statistically significant(P<0.001).There is no statistical difference between the two groups of data on the VDR Ⅱ and VDR Ⅲ fragments.on the statistical results of methylation levels at 33 CpG sites on the VDR Ⅰfragment,the difference between experimental and control groups was statistically significant at CpG-5(p=0.008)and CpG-8(p(0.001)sites.Conclusion:The abnormal methylation level of DNA VDR gene promoter was associated with the occurrence of idiopathic low citrateuria in Bai nationality of Yunnan Province,and its DNA methylation was closely related to the pathogenesis of idiopathic low citrate in SNP locus rs2228570 FF genotype.