蛋鸡胱硫醚β合酶基因启动子鉴定与分析

doi:10.11843/j.issn.0366-6964.2018.01.024

畜牧兽医学报 ›› 2018, Vol. 49 ›› Issue (1): 211-217.doi: 10.11843/j.issn.0366-6964.2018.01.024

蛋鸡胱硫醚β合酶基因启动子鉴定与分析

王涵¹, 陈烨¹, 周荣艳^1,2*, DUNN Ian², 陈辉¹, 锡建中¹, 张振红¹

1. 河北农业大学动物科技学院, 保定 071001;
2. Roslin Institute (Edinburgh), Roslin, Midlothian EH25 9RG, Scotland

收稿日期:2017-07-28 出版日期:2018-01-23 发布日期:2018-01-18
通讯作者: 周荣艳,副教授,E-mail:rongyanzhou@126.com
作者简介:王涵(1992-),男,四川达州人,硕士生,主要从事动物遗传育种与繁殖研究,E-mail:blue-5206@qq.com
基金资助:
河北省首批青年拔尖人才支持计划（2016-2018）；河北农业大学青年学术带头人支持项目（2015-2017）

Identification and Analysis of Promoter of Cystathionine Beta Synthase in Layer

WANG Han¹, CHEN Ye¹, ZHOU Rong-yan^1,2*, DUNN Ian², CHEN Hui¹, XI Jian-zhong¹, ZHANG Zhen-hong¹

1. College of Animal Science and Technology, Hebei Agricultural University, Baoding 071001, China;
2. Roslin Institute(Edinburgh), Roslin, Midlothian EH25 9RG, Edinburgh, UK

Received:2017-07-28 Online:2018-01-23 Published:2018-01-18

摘要/Abstract

摘要：

旨在鉴定胱硫醚β合酶（Cystathionine beta synthase，CBS）基因5'调控区和核心启动子特征，为研究蛋鸡骨骼中该基因的表达调控机制提供依据。采集68周龄罗曼蛋鸡胫骨组织，利用酚-氯仿法提取基因组DNA，以UCSC数据库中CBS基因序列为模板设计5'调控区扩增引物，利用生物信息方法分析5'调控区序列特征并进行转录因子结合位点预测，并构建双荧光素酶报告基因重组载体，通过转染DF-1细胞系后检测双荧光素酶活性，进行CBS基因的核心启动子区鉴定。结果表明，蛋鸡CBS基因5'调控区含有潜在的核心启动子区、典型的CpG岛和多个转录因子Sp1结合位点；不同长度片段均具有启动子活性，但不同片段活性存在显著差异，其中F4片段（-155~+131 bp）具有最高的转录活性，确定该片段为核心启动子区。蛋鸡CBS基因5'调控区结构特征与核心启动子的鉴定和转录因子结合位点的发现为研究该基因在骨骼中的表达调控机制奠定了基础。

关键词: 蛋鸡, 骨骼质量, CBS, 启动子

Abstract:

To provide the theoretical basis for exploring the regulatory mechanism of cystathionine beta synthase(CBS) gene expression in layer bone, the 5' regulatory region and core promoter characteristics were investigated. The tibias were collected from laying hen at 68 weeks old. The genomic DNA was extracted with phenol-chloroform method. The primers for amplifying 5' regulatory region of CBS gene were designed according to the template sequence downloaded from UCSC database. The characteristic of 5' regulatory region and prediction of transcription factors binding sites were analyzed by bioinformatics methods. The constructed dual-luciferase recombinant vectors were transfected into DF-1 cells for detecting luciferase activtity, and the core promoter of CBS gene was identified. The results showed that the potential core promoter, classical CpG island and multiple transcription factor Sp1 binding sites were located in 5' regulatory region of CBS gene. All the fragments had promoter activity, and there were significant difference in promoter activity among different fragments. The fragment F4 (-155-+131 bp) with the highest transcription activity was identified as core promoter region. The identification of core promoter and prediction of transcription factor binding sites of CBS gene provide the basis for studying the expression regulatory mechanism of CBS gene in bone.

Key words: layer, bone quality, CBS, promoter

中图分类号:

S831.2

王涵, 陈烨, 周荣艳, DUNN Ian, 陈辉, 锡建中, 张振红. 蛋鸡胱硫醚β合酶基因启动子鉴定与分析[J]. 畜牧兽医学报, 2018, 49(1): 211-217.

WANG Han, CHEN Ye, ZHOU Rong-yan, DUNN Ian, CHEN Hui, XI Jian-zhong, ZHANG Zhen-hong. Identification and Analysis of Promoter of Cystathionine Beta Synthase in Layer[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2018, 49(1): 211-217.

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蛋鸡胱硫醚β合酶基因启动子鉴定与分析

Identification and Analysis of Promoter of Cystathionine Beta Synthase in Layer

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