[1]肖丹,兰长骏,杨琴,等.近视相关RDH5基因内含子区单核苷酸多态性位点rs3138144（GC）对其启动子功能的影响[J].眼科新进展,2021,41(3):223-226.[doi:10.13389/j.cnki.rao.2021.0046]
　XIAO Dan,LAN Changjun,YANG Qin,et al.Effect of single nucleotide polymorphism locus rs3138144 of myopia-related RDH5 gene on the function of its promoter[J].Recent Advances in Ophthalmology,2021,41(3):223-226.[doi:10.13389/j.cnki.rao.2021.0046]
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近视相关RDH5基因内含子区单核苷酸多态性位点rs3138144（G>C）对其启动子功能的影响/HTML
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《眼科新进展》[ISSN:1003-5141/CN:41-1105/R]

卷:
41卷
期数:
2021年3期
页码:
223-226
栏目:
实验研究
出版日期:
2021-03-05

文章信息/Info

Title:
Effect of single nucleotide polymorphism locus rs3138144 of myopia-related RDH5 gene on the function of its promoter
作者:
肖丹; 兰长骏; 杨琴; 刘阳; 谭青青; 廖萱
637000　四川省南充市，川北医学院附属医院眼科，川北医学院眼视光学系
Author(s):
XIAO Dan; LAN Changjun; YANG Qin; LIU Yang; TAN Qingqing; LIAO Xuan
Department of Ophthalmology,Affiliated Hospital of North Sichuan Medical College, Department of Ophthalmology & Optometry,North Sichuan Medical College,Nanchong 637000,Sichuan Province,China
关键词:
RDH5基因; 单核苷酸多态性; 启动子; 基因表达
Keywords:
RDH5;  single nucleotide polymorphism;  promoter;  gene expression
分类号:
R778
DOI:
10.13389/j.cnki.rao.2021.0046
文献标志码:
A
摘要:
目的　探讨近视相关视黄醇脱氢酶5（RDH5）基因内含子区单核苷酸多态性（SNP）位点rs3138144（G>C）对其启动子功能的影响。方法　从初始密码子ATG往前扩增1500 bp的5’端上游调控区域作为拟研究的启动子片段，以包含rs3138144位点的长220 bp的区段作为研究的内含子片段。与萤火虫荧光素酶报告基因载体pGL3-Basic进行重组质粒的克隆与鉴定，构建rs3138144荧光素酶报告基因质粒野生型（pGL3-RDH5-G）和rs3138144荧光素酶报告基因质粒突变型（pGL3-RDH5-C）载体，同时设定pGL3-Basic作为对照。与内参质粒pRL-TK共转染ARPE-19细胞及HEK293细胞，通过双荧光素酶报告基因系统检测报告基因活性。结果　在ARPE-19细胞中，野生型rs3138144位点荧光素酶相对活性为0.806±0.156，突变型为0.525±0.130，对照为0.085±0.027，野生型荧光素酶相对活性高于突变型及对照，差异均有统计学意义（均为P<0.05）。在HEK293细胞中，野生型rs3138144位点荧光素酶相对活性为0.075±0.013，突变型为0.040±0.006，对照为0.004±0.001,野生型高于突变型及对照，差异均有统计学意义（均为P<0.05）。等位基因G及等位基因C均能与核蛋白中的某种转录因子结合，灰度值分析结果提示等位基因G结合能力更强。结论　RDH5基因内含子区SNP位点rs3138144（G>C）影响RDH5基因启动子转录活性。
Abstract:
Objective　To investigate the effect of single nucleotide polymorphism (SNP) locus rs3138144 (G＞C) in the intron region of myopia-related RDH5 gene on its promoter’s function.Methods　The initial codon ATG forward amplified to 1500 bp in the upstream regulatory region of the 5 ’end was selected as the promoter fragment to be studied, and the 220 bp region containing the rs3138144 locus was used as the intron fragment. The recombinant plasmids were cloned and identified with the firefly luciferase reporter gene vector pGL3-Basic. The wild type (pGL3-RDH5-G) and the mutant type (pGL3-RDH5) of rs3138144 luciferase reporter gene plasmid were constructed, and the pGL3-Basic as a control. ARPE-19 and HEK293 cells were co-transfected with internal reference renilla luciferase reporter gene plasmid pRL-TK, and luciferase activity was detected by double luciferase reporter gene system.Results　In ARPE-19 cells, the relative activity of the luciferase reporter gene at rs3138144 locus was 0.806±0.156 in the wild type, 0.525±0.130 in the mutant type, and 0.085±0.027 in the control. The value of wild type was higher than the mutant type and the control, and the differences were statistically significant (all P<0.05). In HEK293T cells, the relative activity of the luciferase reporter gene at rs3138144 locus was 0.075±0.013 in the wild type, 0.040±0.006 in the mutant type, and 0.004±0.001 in the control. The value of wild type was higher than that in the mutant and the control, and the differences were statistically significant (both P<0.05). Both allele G and allele C can bind to a certain transcription factor in nucleoprotein, and the gray value analysis indicated that the binding ability of allele G was stronger.Conclusion　The SNP site rs3138144 (G＞C) in the intron region can affect the transcription activity of RDH5 gene promoter.

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备注/Memo

备注/Memo:
四川省科技厅应用基础研究项目（编号：2019YJ0381）；博士科研基金项目（编号：CBY20-QD05）
更新日期/Last Update: 2021-03-05