FAM213B基因mRNA和启动子的克隆及序列分析
作者:
作者单位:

作者简介:

通讯作者:

基金项目:

国家自然科学基金(31201771);现代农业产业技术体系专项(CARS-36)


Cloning and sequence analysis of mRNA and promoter of pig FAM213B gene
Author:
Affiliation:

Fund Project:

  • 摘要
  • |
  • 图/表
  • |
  • 访问统计
  • |
  • 参考文献
  • |
  • 相似文献
  • |
  • 引证文献
  • |
  • 资源附件
    摘要:

    【目的】获得猪FAM213B基因完整mRNA和启动子序列,研究猪FAM213B基因表达,为探讨母猪妊娠的建立和胚胎发育调控机制奠定基础。【方法】通过5′RACE和3′RACE技术,获得基因完整mRNA序列,分析不同物种该基因氨基酸序列相似性;通过PCR克隆启动子区,并通过双荧光素酶报告基因载体系统转染猪子宫内膜细胞,研究其转录活性。【结果】猪FAM213B基因mRNA全长808 bp,其中5′UTR、CDS区和3′UTR长度分别为67、 609(含终止密码子)和132 bp(不含poly A序列),在17~106位氨基酸之间存在硫氧还蛋白折叠结构域;与猪FAM213B基因其他2个潜在转录本相比,三者都包含硫氧还蛋白折叠结构域,但蛋白三级结构存在较大差异;猪FAM213B氨基酸序列与山羊、牛和绵羊高度相似,相似性分别为94.03%、 93.03%和91.54%。克隆获得2 261 bp(-2 231/+30)的基因启动子序列,将其连接至双荧光素酶报告基因载体,转染猪子宫内膜细胞,发现获得的启动子片段能够启动下游报告基因的转录,在启动子区存在潜在的典型NFκB等转录因子结合位点。【结论】本研究获得猪FAM213B基因转录本长度为808 bp,其蛋白存在硫氧还蛋白折叠功能结构域,其启动子序列(-2 231/+30)在猪子宫内膜细胞中具有较强的转录活性。

    Abstract:

    【Objective】To obtain the complete mRNA and promoter sequences of pig FAM213B gene, and provide a basis for studying the mechanism of the FAM213B gene in regulating gestation establishment and embryo development of female pigs.【Method】The complete mRNA sequence of the FAM213B gene was obtained using 5′ and 3′RACE methods.The amino acid sequence similarities of different species were analyzed. The gene promoter was cloned, and its transcription activity was detected by the dual luciferase report system in porcine endometrial cells.【Result】The mRNA of pig FAM213Bgene was 808 bp in full length, including the 5′UTR, CDS and 3′UTR of 67, 609 (including the termination codon) and 132 bp (excluding poly A), respectively. A thioredoxin fold domain was predicted from the 17th to 106th amino acid residues. The obtained transcript and the other two computed transcripts all contained the thioredoxin fold domains, but the tertiary structures of three proteins were highly different. The amino acid sequence of pig FAM213B showed 94.03%, 93.03% and 91.54% similarities with those of goat, cattle and sheep, respectively. The cloned promoter sequence (-2 231/+30) of the FAM213B gene was linked with the dual luciferase report vector, and transfected into the endometrial cells. The promoter fragment could drive the expression of the downstream report gene. There were potential binding sites of transcription factors such as NFκB within the promoter. 【Conclusion】The complete sequence of the transcript of pig FAM213B gene is 808 bp. The FAM213B protein contains a thioredoxin fold domain. The FAM213B promoter (-2 231/+30) has strong transcription activity in porcine endometrial cells.

    参考文献
    相似文献
    引证文献
引用本文

张爱玲,孙显月,卢孝璋,李加琪,张豪.猪FAM213B基因mRNA和启动子的克隆及序列分析[J].华南农业大学学报,2017,38(3):1-8

复制
分享
文章指标
  • 点击次数:
  • 下载次数:
历史
  • 收稿日期:2016-07-24
  • 最后修改日期:
  • 录用日期:
  • 在线发布日期: 2017-05-04