红鳍东方鲀SREBP-1和SREBP-2基因克隆及表达分析

doi:10.7668/hbnxb.2016.04.013

摘要/Abstract

摘要： 为了研究红鳍东方鲀SREBP-1和SREBP-2基因的功能及在脂肪代谢中的分子机制，从红鳍东方鲀脂肪组织提取总RNA，反转录获得cDNA模板，利用设计的引物PCR扩增获得SREBP-1和SREBP-2开放阅读框（ORF），SREBP-1基因ORF区的cDNA序列长度为3 330 bp，编码1 109个氨基酸，SREBP-2基因ORF区的cDNA序列为3 444 bp，编码1 147个氨基酸。实时荧光定量PCR检测SREBP-1基因在红鳍东方鲀不同组织中的表达特征，结果表明，SREBP-1在脑组织中表达丰度最高，其次为眼、脂肪组织、肠、鳃、脾脏、心脏及肝脏，在肌肉中表达丰度最低。将SREBP-1连接到原核表达载体pET32a上，利用IPTG对重组质粒pET32a-SREBP-1在大肠杆菌Rosetta （DE3）进行诱导表达，SDS-PAGE电泳显示在141 kDa处有特异性的条带出现。

关键词: 红鳍东方鲀, SREBP-1基因, SREBP-2基因, 原核表达

Abstract: In order to study the function and molecular mechanism of SREBP-1 and SREBP-2, the open reading frames (ORF) of torafugu SREBP-1 and SREBP-2 genes were amplified from total RNA of adipose tissue by reverse transcription-polymerase chain reaction (RT-PCR).The results showed that the ORF of SREBP-1 was composed of 3 330 bp,encoding 1 109 amino acids,while the ORF of SREBP-2 was composed of 3 444 bp,encoding 1 147 amino acids.The expression of SREBP-1 gene in torafugu tissues was determined using Real-time quantitative PCR,presenting that the highest expression was in brain,followed in eye,adipose tissue,spleen,gill,intestine,heart and liver,but the lowest in muscle.The prokaryotic expression system of recombined vector pET32a-SREBP-1 was constructed successfully.SDS-PAGE analysis showed that the expressed protein accumulated and the molecular weight of expressed fusion protein was 141 kDa.

Key words: Takifugu rubripes, SREBP-1 gene, SREBP-2 gene, Prokaryotic expression

中图分类号:

S917
Q786

张伟, 姬明丽, 郭前进, 千智斌. 红鳍东方鲀SREBP-1和SREBP-2基因克隆及表达分析[J]. 华北农学报, 2016, 31(4): 74-79. doi: 10.7668/hbnxb.2016.04.013.

ZHANG Wei, JI Mingli, GUO Qianjin, QIAN Zhibin. Cloning and Expression of SREBP-1 andSREBP-2 in Torafugu Takifugu rubripes[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2016, 31(4): 74-79. doi: 10.7668/hbnxb.2016.04.013.

http://www.hbnxb.net/CN/Y2016/V31/I4/74

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