遗传 ›› 2009, Vol. 31 ›› Issue (11): 1107-1112.doi: 10.3724/SP.J.1005.2009.01107

• 研究报告 • 上一篇    下一篇

绵羊CAST基因2型和4型转录本的克隆及特性分析

张菊, 杜立新, 魏彩虹, 李宏滨

  

  1. 中国农业科学院北京畜牧兽医研究所 国家畜禽分子遗传育种中心, 北京 100193

  • 收稿日期:2009-04-19 修回日期:2009-09-20 出版日期:2009-11-10 发布日期:2009-11-10
  • 通讯作者: 杜立新

Cloning and characterization of CAST transcript 2 and 4 in sheep

ZHANG Ju, DU Li-Xin, WEI Cai-Hong, LI Hong-Bin   

  1. National Center for Molecular Genetics and Breeding of Animal, Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing 100193, China
  • Received:2009-04-19 Revised:2009-09-20 Online:2009-11-10 Published:2009-11-10
  • Contact: DU Li-Xin

摘要:

钙蛋白酶抑制蛋白(Calpastatin, CAST)是一种内源性的需要Ca2+激活的钙蛋白酶抑制剂, 在肌肉组织的蛋白质降解过程中起重要的调节作用。文章利用牛CAST基因的mRNA序列, 通过逆转录RT-PCR首次克隆获得绵羊CAST基因2型转录本和4型转录本的部分cDNA序列, 并对序列进行生物信息学分析。CAST基因2型转录本的扩增片段为4 385 bp, 完整的开放阅读框为2 361 bp, 编码786个氨基酸; CAST基因4型转录本的扩增片段为1 467 bp, 完整的开放阅读框为1 317 bp, 编码438个氨基酸。CASTⅡ型蛋白序列存在4个保守结构域, CASTⅣ型蛋白序列存在3个保守结构域; 两者的二级结构均以螺旋为主, 富含疏水区域, 其氨基酸序列存在多个磷酸化位点以及蛋白激酶C(Protein kinase C, PKC)的磷酸化位点。通过RT-PCR分析CAST基因2型转录本和4型转录本的组织表达谱, 结果表明CAST基因2型转录本在所检测的10个组织中均表达, CAST基因4型转录本仅在睾丸组织中表达。

关键词: cDNA克隆, CAST, 组织表达

Abstract:

As an endogenous inhibitor of the calpain system activated by Ca2+, calpastatin (CAST) plays a regulatory role in muscle proteolysis. Based on the bovine mRNA sequences, part of cDNA fragments of sheep CAST transcript 2 and 4 were obtained by RT-PCR. Bioinformatic analysis showed that sheep CAST transcript 2 was 4 358 bp in length with an open reading frame (ORF) 2 361 bp long and encoded 786 amino acids, while sheep CAST transcript 4 was 1 467 bp in length with 1 317 bp ORF encoding 438 amino acids. It was predicted that CAST type II contained four conserved domains and CAST type IV contained three conserved domains, and their secondary structures were rich in both hydrophobic regions and helical regions, with certain conserved phosphorylation sites and phosphorylation sites of protein kinase C (PKC). RT-PCR was conducted to analyze the expression patterns of CAST transcript 2 and transcript 4. CAST transcript 2 was expressed in ten tissues detected while CAST transcript 4 only in testis.