猪StAR基因启动子区克隆及其转录活性研究

doi:10.11843/j.issn.0366-6964.2018.07.023

Abstract

Abstract:

The research was designed to analyze the activity region in the promoter of StAR gene, and to explore the mechanism of expression regulation of StAR gene, thus provide new ideas for improving the fertility of the pigs from the perspective of breeding. The sequence of the promoter activity region was analyzed by online tools based on the 5'-flanking sequence of swine StAR gene published by Ensembl database. The specific primer was designed and the PCR was used to amplify the gene promoter sequence based on the reference genomic sequence of the pigs, pGL3-StAR promoter luciferase reporter gene vectors were constructed and transfected into 293T cells, the relative luciferase activity was measured by dual luciferase assay system. The results showed that the 5'-flanking sequence of swine StAR gene didn't contain the typical TATA-box and CpG island. Ten promoter fragments with different lengths were obtained and luciferase reporter gene vectors were constructed, and then to analyze their transcriptional activity through transfected into 293T cells, respectively. The core promoter region of StAR gene was located in the 5'-flanking sequence of swine StAR gene, among them, -196-+127 bp region had the highest activity value, which was significantly higher than other deletion fragments(P<0.01),suggested that the region of +127——196 bp existed an important positive regulatory element. The exon 1 played a critical role in regulating the activity of the promoter. -41——196 bp as the core promoter region, contained a primary positive regulatory element, in which there were a number of transcription factor binding sites, including GATA2, GATA4, SP1, ZNF263, Hoxa9, KLF16 and ZNF740. Bioinformatics analysis of StAR gene and detection of dual reporter gene activity with different length promoter fragments confirmed that 5'-flanking sequence of swine StAR had promoter transcriptional activity. The promoter region of the gene had been preliminarily identified, the promoter region and the main regulatory region were found, which provided a theoretical basis for further study of StAR gene transcriptional regulation mechanism.

Key words: StAR gene, promoter, luciferase activity, reporter gene

CLC Number:

S828.2
Q785

HU Hui-yan, JIA Qing, HOU Sheng-kui, LIU Jin, ZHANG Jing, ZHANG Wei-feng, XI Jian-zhong. Cloning and Transcriptional Activity of Swine StAR Gene Promoter[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2018, 49(7): 1524-1532.

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Cloning and Transcriptional Activity of Swine StAR Gene Promoter

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