CART基因启动子双荧光素酶重组载体构建

Construction of CART Gene Promoter Double Luciferase Recombinant Vector

可卡因—苯丙胺调节转录肽(cocaine-and amphetamine regulated transcript peptides,CART)是一种内源性神经肽,广泛分布于中枢神经和外周神经系统。CART具有多种不同的生理功能,在牛繁殖机能研究中发现,CART还可以通过影响雌激素分泌抑制卵泡发育。这一发现为深入分析牛卵泡闭锁以及提高母畜繁殖率提供理论依据。然而%CART%基因转录调控机制尚不明确。研究利用pGL3系列萤火虫荧光素酶表达载体,%Mlu%I、%Xho%I双酶切割牛%CART%基因启动子-475 bp~+22 bp区域,成功构建重组质粒,转染293T细胞,检测相对荧光活性,为后续筛选%CART%基因核心启动子活性片段创造条件,进而为%CART%转录调控分子机制的探究奠定基础。

Cocaine-and amphetamine regulated transcript peptides(CART)is an endogenous neuropeptide widely distributed in the central and peripheral nervous systems.CART has a variety of different physiological functions.In the study of bovine reproductive function,it has been found that CART can also inhibit follicle development by affecting estrogen secretion.This finding provides a theoretical basis for further analysis of bovine follicular atresia and improving the reproductive rate of female animals.However,the mechanism of CART gene transcriptional regulation remains unclear.In this study,pGL3 series firefly luciferase expression vector was used to cut the-475 bp~+22 bp region of bovine CART gene promoter with Mlu I and Xho I double enzymes.The recombinant plasmid was successfully constructed and transfected into 293T cells to detect the relative fluorescence activity,which created conditions for subsequent screening of the active fragment of CART gene core promoter.Thus,it lays a foundation for exploring the molecular mechanism of CART transcriptional regulation.