中国农业科学 ›› 2013, Vol. 46 ›› Issue (4): 830-840.doi: 10.3864/j.issn.0578-1752.2013.04.017

• 畜牧·兽医·资源昆虫 • 上一篇    下一篇

鸭血清白蛋白基因克隆及其在病毒感染过程中的表达变化

 徐琪, 陈阳, 李秀, 黄正洋, 张扬, 李欣钰, 童一宇, 段修军, 陈国宏   

  1. 1.江苏省动物遗传繁育与分子设计重点实验室,江苏扬州 225009
    2.江西农业大学动物科学技术学院,南昌 330045
    3.国家水禽种质资源基因库,江苏泰州 225300
  • 收稿日期:2012-07-06 出版日期:2013-02-15 发布日期:2012-09-05
  • 联系方式: 徐琪,E-mail:xuqi@yzu.edu.cn
  • 基金资助:

    国家自然科学基金(31101704)、现代农业产业技术体系建设专项CARS-43-3)、江苏省属高校自然科学基础研究面上项目(07KJB230138)江苏高校优势学科建设工程资助项目(苏政办发【2011】137号)

Molecular Cloning and Expression Pattern of Duck Serum Albumin Gene During Viral Infection

 XU  Qi, CHEN  Yang, LI  Xiu, HUANG  Zheng-Yang, ZHANG  Yang, LI  Xin-Yu, TONG  Yi-Yu, DUAN  Xiu-Jun, CHEN  Guo-Hong   

  1. 1.Jiangsu Key Laboratory of Animal Genetics, Breeding and Molecular Design, Yangzhou 225009, Jiangsu;
    2.School of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045;
    3.National Waterfowl Germplasm Resorurse,    Taizhou 225300, Jiangsu
  • Received:2012-07-06 Published:2013-02-15 Online:2012-09-05

摘要: 【目的】克隆鸭血清白蛋白(duck serum albumin,DSA)基因,并对其进行生物信息学分析和mRNA表达规律研究。【方法】以前期抑制性消减杂交技术筛选的白蛋白(albumin,ALB)基因为候选基因,通过构建雏鸭肝炎病毒和聚肌苷酸胞苷酸(polyinosinic polycytidylic acid,Poly(I:C))感染模型,利用RT-PCR、RACE技术和基因组步移技术分别克隆ALB基因cDNA序列和5′侧翼序列,并对其进行生物信息学分析;同时利用RT-qPCR检测ALB基因各组织时空表达量。【结果】①ALB cDNA全序列长为2 107 bp,包括47 bp的 5′UTR、212 bp的 3′UTR和1 848 bp的开放阅读框(open reading frame,ORF);其5′侧翼序列具有典型的TAAT box、CAAT box以及HSF、HNF、C/EBP等多个肝脏富含的潜在转录因子结合位点;②RT-qPCR显示,ALB mRNA 呈肝脏组织特异性表达,且在雏鸭肝炎病毒和Poly(I:C)等抗原刺激后,肝脏组织ALB mRNA表达量总体表现水平为先上升后下降,24 h后保持在稳定水平。【结论】成功克隆了鸭ALB基因cDNA和5′侧翼序列,该基因在不同禽类(鸡、鸭、火鸡)中表现为相当保守,主要在肝脏组织中表达,且在雏鸭肝炎病毒和Poly(I:C)感染下,ALB mRNA表现水平为先上升后下降。

关键词: 鸭 , ALB基因 , 克隆 , 基因表达

Abstract: 【Objective】This experiment was conducted to clone serum albumin (ALB) gene and explore albumin gene molecular structure and expression regularity in duck. 【Method】Screening albumin as a candidate gene based on the results of suppression subtractive hybridization (SSH). The disease models infected duckling virus hepatitis and polyriboinosinic polyribocytidylic acid, Poly(I:C) were constructed. The cDNA and 5′ flanking sequence of ALB were obtained by RT-PCR, RACE and genome walking, and then the sequences were analyzed by bioinformatics approaches. The temporal spatial expression of ALB in tissues was detected by RT-qPCR. The results showed that the full-length cDNA of ALB gene was 2 107 bp, including 47 bp of 5′UTR, 212 bp of 3′UTR, and 1 848 bp of open reading frame (ORF). And the potential transcriptional factor binding sites were predicted in 5′ flanking region including TAAT box, CAAT box, HSF, HNF, C/EBP and so on in liver. The results of RT-qPCR showed that ALB mRNA was specifically expressed in liver, the ALB mRNA levels of livers decreased first, then increased and remained stable 24 hours late in the 3-day-old ducklings treated with duckling virus hepatitis and Poly(I:C). 【Conclusion】The cDNA and 5′ flanking sequence of ALB were detected successfully in duck, its genetic relationship with other kinds of flow were closing by constructing phylogenetic tree. The albumin gene expression regularity was discovered after infected duckling virus hepatitis and Poly(I:C).

Key words: duck , albumin gene , molecular cloning , gene expression