中国畜牧兽医 ›› 2014, Vol. 41 ›› Issue (9): 25-31.

• 生物技术 • 上一篇    下一篇

快长系F1鹅主要组织相容性复合物Ⅰ基因的克隆与序列分析

杨丽金1, 朱峰伟1,2, 吕春伟1, 朱新产1   

  1. 1. 青岛农业大学生命科学学院, 山东省高校植物生物技术重点实验室, 山东青岛 266109;
    2. 青岛即墨市畜牧兽医局, 山东即墨 266200
  • 收稿日期:2014-01-22 出版日期:2014-09-20 发布日期:2014-09-24
  • 通讯作者: 朱新产 E-mail:xczhu321@126.com
  • 作者简介:杨丽金(1990- ),女,山东人,硕士,研究方向:病毒感染致病机制。
  • 基金资助:

    山东省自然基金(ZR2011CM008);青岛市科技基金资助项目(12-1-4-5-(2)-jch)。

Cloning and Sequence Analysis of Major Histocompatibility Complex Ⅰ Gene from Goose F1 of Fast-growth Lines

YANG Li-jin1, ZHU Feng-wei1,2, LV Chun-wei1, ZHU Xin-chan1   

  1. 1. Key Laboratory of Plant Biotechnology in University of Shandong Province, College of Life Science, Qingdao Agricultural University, Qingdao 266109, China;
    2. Jimo Bureau of Animal Husbandry and Veterinary, Jimo 266200, China
  • Received:2014-01-22 Online:2014-09-20 Published:2014-09-24

摘要: 根据GenBank中已发表的鸭科(Anatidae)雁属水禽主要组织相容性复合物Ⅰ(major histocompatibility complex Ⅰ,MHC Ⅰ)基因的多重比对确定保守序列,用Primer Premier 5.0软件在保守序列区域设计1对特异性引物,从快长系F1鹅的基因组DNA中扩增获得MHCⅠ基因片段序列,采用DNAsis、Mega 5.10、BLAST等多种生物学软件对核苷酸序列和氨基酸序列的结构、性质和功能进行分析。结果显示,克隆的F1鹅MHCⅠ基因片段长1036 bp,编码96个氨基酸,与NCBI中五龙鹅MHCⅠ基因和编码序列的同源性达93%和83%,有72处碱基序列不同,16个氨基酸发生改变。与其他家禽也有较高的同源性,亲缘关系依次为四季鹅>鸡>鸭。F1鹅MHCⅠ基因片段编码蛋白分子质量为11.342 ku,PI值为5.32,不稳定系数为34.92,脂肪系数为42.81,平均疏水性为-1.066,可能存在9个B细胞抗原表位,但不含信号肽,提示该蛋白为亲水性的非分泌蛋白,具有较高的免疫原性。在蛋白的二级结构中α-螺旋占31.25%,β-折叠占16.67%,β-转角占14.58%,无规则卷曲占37.50%,三级结构呈现近似哑铃型,具有2个结构域:氨基末端结构域和羧基末端结构域。因此,环境中的病原压力使MHC基因在物种之间和种群的个体之间都产生了明显差异,存在着多态性MHC分子与多样性抗原肽相互作用的关系。MHC决定着疾病易感性个体的差异,是研究疾病抗性的最佳候选标记基因。

关键词: 快长系F1鹅; 主要组织相容性复合物Ⅰ基因; 克隆; 生物信息学

Abstract: According to the multiple alignments identified major histocompatibility complex Ⅰ (MHC Ⅰ) gene conserved sequence registered in GenBank from the family ducks (Anatidae) anser waterfowl, a pairs of specific primers for the fragments of MHCⅠgene of goose F1 from fast-growth lines were designed and synthesized by Primer Premier 5.0. Using the genome DNA of goose F1 from fast-growth lines, the target gene fragment was obtained by PCR. To conduct sequencing of the fragments of MHCⅠgene of goose F1 from fast-growth lines and make sequence alignment and analysis of protein structure and function by bioinformatics, and research the characteristics of MHCⅠgene of goose F1 and the physicochemical properties of the protein. Bioinformatics was analyzed the nucleic acid data, deduced amino acid sequence and phylogenetic trees. The result of sequence analysis showed that the fragments of MHCⅠgene of goose F1 from fast-growth lines was 1036 bp in length, which coded 96 amino acids polyprotein. The homology were 93% and 83% with MHC Ⅰ gene and coding sequence of Wulong goose in NCBI respectively. There were 72 different bases sequence and 16 amino acids change. There also was higher homology with other poultry, and existed genetic relationship of Siji goose > chickens > ducks.The homology segment sequences corresponding to the fragments of MHCⅠ gene of goose F1 coded 96 amino acids protein, which molecular weight, PI, positively or negatively charged amino acid, estimated half-life, instability index, aliphatic index and average hydrophobicity were 11.342 ku, 5.32, 14, 17, 2.8 h, 34.92, 42.81, -1.066, respectively, and appeared 9 B cell epitopes, but contained no signal peptide. These results indicated that the protein for hydrophilic non-secreted proteins, had the high immunogenicity. In addition, The protein structure study indicated that alpha-helix, beta-sheet, beta-turn and random coil were 31.25%,16.67%, 14.58% and 37.50%, respectively. There existed amino terminal domain and carboxyl terminal domain in the tertiary structure. Therefore, MHC gene had significant difference between species and populations of individuals by the pathogen pressure in environment, and there were the interaction between polymorphism of MHC molecules and the diversity of antigenic peptide. MHC determined the differences of individual susceptibility to disease, and could be treated as a candidate gene for disease resistance.

Key words: goose F1 of fast-growth lines; MHCⅠ gene; cloning; bioinformatics

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