Prss37基因转录起始位点的确定及其转录调控的初步探讨  

Identification of Mouse Prss37 Transcription Initiation Site and Preliminary Analysis of Its Transcription Regulation

作　　者：王津津 匡昉哲 庄华 陆翠杰 匡颖 

机构地区：[1]上海南方模式生物科技发展有限公司,上海201203 [2]上海南方模式生物研究中心,上海201203 [3]同济大学生命科学与技术学院,上海200092

出　　处：《实验动物与比较医学》2016年第2期93-100,共8页Laboratory Animal and Comparative Medicine

基　　金：上海市科委项目资助（13140901400,14140900400,13DZ2280600）

摘　　要：目的确定P坶537基因的转录起始位点，并利用转基因技术在整体水平验证该启动子的组织特异性。方法运用cDNA5’末端快速扩增法（5’RACE）对C57BL／6J小鼠睾丸mRNA进行分析，确定P坶s37的转录起始位点；构建P船s37内源启动子区域转录起始位点上游不同长度DNA片段（1kb、2kb、4kb）驱动的荧光素酶基因表达的质粒；通过受精卵雄原核显微注射技术获得上述三种启动子驱动荧光素酶基因表达的转基因小鼠；利用活体成像技术观察荧光素酶基因的表达，明确启动子的组织特异性。结果Prss37基因的转录起始位点位于NCBIGeneBankTM（NM_026317．2）报道序列上游的40bp处；成功获得三种转基因小鼠，其中1kb启动子驱动的转基因小鼠检测到荧光素酶基因的表达，且在脑、睾丸和附睾组织中的表达较强。结论明确P瑙妇7的转录起始位点，成功获得了睾丸和附睾表达荧光素酶基因的转基因小鼠，为进一步的Prss37基因转录调控研究奠定了基础。Objective To determine the transcription initiation site of mouse Prss37 gene, and verify the tissue specificity of the promoter in vivo using transgenic technology. Methods The transcription initiation site was identified by 5＇ rapid amplification of cDNA ends （5＇ RACE ） used to analyze the mRNA of testis from C57BL/6J mice. The plasmids including different lengths of promoter （lkb, 4kb, and 6kb） and luciferase gene were constructed. Then, these plasmids were injected into male pronucleus by microinjection technique to obtain luciferase positive transgenic mice. Results The transcription start site of Prss37 gene was located at the 40bp site upstream of the predicted site reported in GeneBankTM （NM_026317.2）. Moreover, three kinds of transgenic mice were successfully established, but the expression of luciferase gene was only detectable in transgenic mice driven by lkb promoter. The higher expression level of luciferase was found in brain, testis and epididymis tissues. Conclusion The transgenic mice expressing luciferase in testis and epididymis tissues were successfully established, which laid a foundation for further research on the transcriptional regulation of Prss37 gene.

关 键 词：Prss37基因 转录 转基因小鼠 荧光素酶基因 

分 类 号：Q75[生物学—分子生物学]