承德无角山羊KCNJ15基因核心启动子区鉴定
Identification of the Core Promoter Region of KCNJ15 Gene in Chengde Polled Goat(Capra hircus)
张文浩 1刘爽 1韩聪 1洪支林 1谢遇春 1乔贤 1刘铮铸 1巩元芳1
作者信息
- 1. 河北科技师范学院动物科技学院/河北省特色动物种子资源挖掘与创新重点实验室,秦皇岛 066004
- 折叠
摘要
角是山羊(Capra hircus)的重要表型性状,KCNJ15(potassium inwardly rectifying channel subfamily J member 15)是影响山羊角生长的重要基因之一.本研究以承德无角山羊为实验动物,利用PCR技术扩增KCNJ15基因2 353 bp长度序列(起始密码子上游2064 bp～下游289 bp),通过在线软件预测核心启动子区、转录因子结合位点和调控元件;进一步构建不同缺失片段的启动子报告基因载体,瞬时转染293T细胞,使用双荧光素酶检测试剂盒检测各缺失片段的活性,确定KCNJ15基因的核心启动子区.结果显示,成功获得承德无角山羊KCNJ15基因(2353 bp)序列,软件预测-423～+289 bp可能为启动子活性区,同时预测到多个转录因子结合位点以及CAAT和TATA调控元件.双荧光素酶活性检测结果显示,在4个启动子缺失片段中,-491～+289 bp的相对活性值最高,提示该区域是核心启动子区,与软件预测结果基本一致.本研究为深入探究KCNJ15基因调控山羊角性状分子机制提供了基础资料.
Abstract
Horn is an important trait in goats(Capra hircus),and the potassium inwardly rectifying channel subfamily J member 15(KCNJ15)gene is an important gene,which affects the growth of horn.The present study selected Chengde polled goat as the test animal and extracted genomic DNA.Then the 2353 bp sequence(2064 bp upstream～289 bp downstream of the start codon)of KCNJ15 gene of Chengde polled goat was amplified by PCR.The sequence was analyzed by online software,the plasmids with different deletion fragments were constructed and transiently transfected into 293T cells.In order to determine the core promoter region of the KCNJ15 gene,dual luciferase detection kit was used to measure the activity.The sequence of Chengde polled goat(2353 bp)was successfully obtained,and software prediction showed that-423～+289 bp might be the active region in the promoter.At the same time,several transcription factor binding sites and CAAT and TATA regulatory elements were simultaneously predicted.The dual luciferase activity assay showed that-491～+289 bp had the highest relative activity value among the 4 fragments,suggesting that this region might be the core promoter region,which was consistent with the results predicted by software.This study provides basic data for in-depth exploration of the molecular regulation mechanism of KCNJ15 gene in goat horn traits.
关键词
承德无角山羊/KCNJ15(potassium/inwardly/rectifying/channel/subfamily/J/member/15)基因/核心启动子区Key words
Chengde polled goat/Potassium inwardly rectifying channel subfamily J member 15(KCNJ15)gene/Core promoter region引用本文复制引用
基金项目
河北省羊产业技术体系创新团队遗传资源开发与利用岗项目(HBCT2023200201)
出版年
2024
国家科技期刊平台
NSTL
万方数据