精子顶体相关基因SPACA7的转录活性研究

Study on transcription activity of sperm acrosome related gene SPACA7

目的研究精子顶体相关基因SPACA7转录起始位点上游-2 000bp至下游+84bp范围内启动子活性。方法利用启动子截短实验获得人类SPACA7基因转录起始位点上游-2 000bp与下游+84bp序列的系列不同长度的启动子共16个,以人肾上皮细胞293T基因组DNA为模板,进行PCR反应,将不同长度的启动子重组至荧光素酶表达载体pGL3-basic中并转染细胞,转染24h后收集细胞加入裂解液,收集细胞裂解液,利用多功能酶标仪测定荧光素酶活性。结果我们研究了SPACA7启动子-2 000bp至+84bp序列,构建了16个不同长度的promoter::Luciferase,荧光素酶活性检测结果发现各个启动子都具有一定的活性,其中-1 500bp至+84bp序列启动子活性最高。结论初步证明精子顶体相关基因SPACA7启动子在-1 500bp至+84bp区域具有较强转录活性。

Objective：To investigate the promoter activity in the region from-2000 bp upstream to＋84 bp downstream of the transcription initiation site of sperm acrosome related gene SPACA7.Methods：A total of 16 promoters of different lengths in the sequence of-2000 bp upstream and 84 bp downstream of the transcription start site of human SPACA7 gene were obtained by promoter truncation experiments.The 293 Tgenomic DNAs of human kidney epithelial cells were used as a template for PCR.The different lengths of promoters were recombined into luciferase expression vector pGL3-basic and transfected cells.After 24 hours of transfection,the cells were collected and lysates were added,and then the cell lysates were collected.The luciferase activity was measured by using a multi-functional microplate reader.Results：The SPACA7 promoter from-2000 bp to＋84 bp sequence was investigated,and 16 different length of the promoters：：luciferase were constructed.Luciferase activity test results found that each promoter has certain activity;in which-1500 bp to＋84 bp sequence promoter activity is the highest.Conclusions：It was initially proved that the sperm acrosome-related gene SPACA7 promoter has strong transcriptional activity in the-1500 bp to＋84 bp region.