引用本文:	黄锦炉,汪开毓,肖丹,王均,付希,王浩丞,连海.罗非鱼源无乳链球菌(Streptococcus agalactiae)Sip基因的克隆、鉴定及分子特性分析.海洋与湖沼,2011,42(4):554-560.

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*罗非鱼源无乳链球菌(Streptococcus agalactiae)Sip基因的克隆、鉴定及分子特性分析*
黄锦炉^1,2, 汪开毓^1,2, 肖丹³, 王均^1,2, 付希^1,2, 王浩丞^1,2, 连海²
1.四川农业大学鱼病研究中心;2.动物疫病与人类健康四川省重点实验室四川农业大学;3.通威股份有限公司

摘要:

利用设计的特异性引物, 扩增出了分离自患病罗非鱼无乳链球菌强毒株 Sip 基因, 并将其克隆到pMD19-T载体上, 之后对重组质粒进行了PCR和双酶切(BamHⅠ+ HindⅢ)鉴定, 并利用多种生物信息学软件对 Sip 基因进行分子特性分析。结果显示, 罗非鱼源无乳链球菌 Sip 编码氨基酸序列具有极高保守性, 与人源、哺乳动物源无乳链球菌亲缘性达 100%, 存在 1 个由 25 个氨基酸组成的信号肽, 具有1个与免疫调节功能相关的LysM结构域, 具有与蛋白翻译后修饰功能相关的磷酸化位点 33 个和 N-糖基化位点 2 个, 编码多肽链中疏水区大于亲水区, 是一种膜外蛋白。密码子偏爱性分析表明, 罗非鱼源无乳链球菌 Sip 基因密码子使用频率差异较大, 密码子偏爱性与真核生物较为接近。

关键词: 无乳链球菌, Sip 基因, 克隆, 分子特性

DOI：10.11693/hyhz201104014014

分类号:

基金项目:教育部“长江学者和创新团队发展计划”创新团队项目, IRT0848 号; 通威股份有限公司重点资助项目, 2006—2009

附件

CHARACTERIZATION OF THE Sip GENE IN THE STRAIN STREPTOCOCCUS AGALACTIAE ISOLATED FROM TILAPIA

HUANG Jin-Lu^1,2, WANG Kai-Yu^1,2, XIAO Dan², WANG Jun^1,2, FU Xi^1,2, WANG Hao-Cheng^1,2, LIAN Hai³

1.Fish Disease Research Center of Fisheries Department of Sichuan Agricultural University;2.Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University;3.Tongwei Co. Ltd.

Abstract:

The Sip gene of a virulent strain of Streptococcus agalactiae isolated from Tilapia was amplified using the PCR method with specific primers and then cloned into pMD19-T vector. The presence of the recombinant plasmid was confirmed by the PCR method combined with the restriction enzyme digestion method (BamHⅠand HindⅢ). The results showed that the amino acid sequence derived from the Sip of S. agalactiae in Tilapia was highly conserved and had a 100% of homology among strains isolated from humans and other mammals. The polypeptide contained a signal peptide consisting of 25 amino acids and a LysM functional domain which was related to immunoregulation. The polypeptide had a number of important sites related to post-translational modification, including 33 phosphorylation sites and 2 N-glycosylation sites. The hydrophobic regions of the Sip were larger than the hydrophilic regions and the Sip was predicted to be located outside of the cell membrane. An analysis of codon bias demonstrated that the codon usage frequency of Sip in the strain S. agalactiae was distinctly different and it resembles more closely to those of the eukaryotes.

Key words: Streptococcus agalactiae, Sip gene, Clone, Molecular characteristics