科尔沁牛乳腺炎临床分离无乳链球菌菌毛岛屿PI-2A辅助蛋白2基因的克隆及序列分析
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- 英文论文题名：Cloning and sequencing of PI-2a ancillary protein 2 gene of bovine mastitis Streptococcus agalactiae
- 论文作者：锡林高娃 ; 杜长智 ; 吴金花 ; 于辰龙 ; 布日额
- 英文论文作者：XI Lin-gao-wa ; DU Chang-zhi ; WU Jin-hua ; WANg Hua ; BAI Wen-li ; YU Chen-long ; BU Ri-e ; Research Institute of Pathogenic in Milk ; College of Life Science ; Inner Mongolia University for Nationalities
- 年：2016
- 作者机构：内蒙古民族大学生命科学学院;内蒙古民族大学乳源性致病菌研究所;
- 论文关键词：乳腺炎 ; 牛 ; 无乳链球菌 ; 菌毛岛屿 ; AP2基因 ; 克隆 ; 序列分析
- 英文论文关键词：Pilus island ; AP2 genes ; Cloning ; Sequencing
- 会议召开时间：2016-07-20
- 会议录名称：中国畜牧兽医学会兽医公共卫生学分会第五次学术研讨会论文集
- 语种：中文
- 分类号：S858.23
- 学会代码：SYGW
- 会议名称：中国畜牧兽医学会兽医公共卫生学分会第五次学术研讨会
- 会议地点：中国内蒙古通辽
- 主办单位：中国畜牧兽医学会兽医公共卫生学分会
- 学会名称：中国畜牧兽医学会兽医公共卫生学分会
- 页数：8
- 文件大小：681k
- 原文格式：D
- 会议级别：全国
摘要
目的克隆内蒙古东部地区科尔沁牛乳腺炎无乳链球菌菌毛岛屿PI-2A辅助蛋白2(AP2)基因片段,并进行序列分析。方法根据Gen Bank中登录的牛乳腺炎无乳链球菌菌毛岛屿PI-2a辅助蛋白2基因序列,使用生物信息学软件Primer5.0设计1对特异性引物,以内蒙古东部地区乳源性无乳链球菌临床菌株总DNA为模板,扩增AP2基因片段,与p MD18-T Vector连接,转化至感受态E.coli JM109中,提取质粒,进行PCR及单、双酶切鉴定;将质粒p MD18-T-AP2转化感受态E.coli JM109后,利用DNAStar分析测序结果,并对其推导的氨基酸序列进行抗原可及性分析,Prot Scale在线程序分析其疏(亲)水性。结果质粒p MD18-T-AP2经PCR及单、双酶切鉴定,证明构建正确;扩增的AP2基因序列长度为699 bp,编码233个氨基酸残基,与Gen Bank中登录的无乳链球菌菌毛岛屿PI-2a的AP2基因核苷酸序列相似性达99%,有3个碱基出现变异(402:C-A;563:C-T;467:A-T),氨基酸序列有两处出现变异,分别为178氨基酸(V-A)和216氨基酸(E-V);该序列二级结构α折叠分布C-末端较多,β折叠较丰富,分布较均匀,无规则卷曲分布在两端,亲水性指数相对较高,抗原性较好。结论成功克隆出内蒙古东部地区牛乳腺炎无乳链球菌菌毛岛屿PI-2a辅助蛋白AP2基因,为进一步研究AP2基因编码蛋白的抗原性奠定了基础。
Purpose Cloning of bovine mastitis Streptococcus agalactiae PI-2a island accessory protein gene AP2,and sequencing.Methods According to the published PI-2a fimbriae island AP2 accessory protein gene sequences of bovine Streptococcus agalactiae on Gen Bank.A pairs of specific primers was designed with bioinformatics software of Primer5.0,and amplification of AP2 gene sequences with the Streptococcus agalactiae total DNA as the template of the Streptococcus agalactiae clinical strains form the Eastern Inner Mongolia.Result The amplified AP2 gene sequences had length 699 bp.and had 99% of similarity to AP2 gene sequences with the bovine Streptococcus agalactiae AP2 gene(EU930008) published on Gen Bank.Conclusion The result showed that successfully cloned the AP2 gene of the bovine mastitis Streptococcus agalactiae of the eastern part of Inner Mongolia of China,and laid the foundation for further study on the antigenicity of the AP2 gene coding protein.
Purpose Cloning of bovine mastitis Streptococcus agalactiae PI-2a island accessory protein gene AP2,and sequencing.Methods According to the published PI-2a fimbriae island AP2 accessory protein gene sequences of bovine Streptococcus agalactiae on Gen Bank.A pairs of specific primers was designed with bioinformatics software of Primer5.0,and amplification of AP2 gene sequences with the Streptococcus agalactiae total DNA as the template of the Streptococcus agalactiae clinical strains form the Eastern Inner Mongolia.Result The amplified AP2 gene sequences had length 699 bp.and had 99% of similarity to AP2 gene sequences with the bovine Streptococcus agalactiae AP2 gene(EU930008) published on Gen Bank.Conclusion The result showed that successfully cloned the AP2 gene of the bovine mastitis Streptococcus agalactiae of the eastern part of Inner Mongolia of China,and laid the foundation for further study on the antigenicity of the AP2 gene coding protein.
引文
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[8]布日额,任晓峰,吴金花,等.牛乳腺炎无乳链球菌表面蛋白pgk抗原优势区的原核表达及其间接ELISA检测方法的建立[J].中国预防兽医学报,2013,35(4);312-6
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[10]郎景民,布日额,吴金花,等.牛乳腺炎无乳链球菌内蒙古分离株sip基因抗原表位序列的高效表达及其抗原性鉴定[J].中国病原微生物学杂志,2010,5(11);805-9.
[11]Telford JL,Barocchi MA,Margarit I,et al.Pili in Gram-positive pathogens[J].Nat Rev Microbiol,2006,4(7):509-19.
[12]Brintoncc,Bryan J,Dillon J,et al.Uses of pili in gonorrhea control;role of bacterial pili in disease,purification and properties of gonococcal pili,and progress in the development of a gonococcal pilus vaccine for gonorrhea.In Brooks G,Gotschlich EC,Holmes KK,Sawyer S and Young F(ed.)Immunobiology of Neisseria gonorrhoeae[M].American Society for Microbiology,1978.155-78.
[2]吴金花.通辽地区奶牛乳房炎主要病原菌分离及药物敏感性试验[J].黑龙江畜牧兽医,2007,3;7-8.
[3]王桂琴,吴聪明,宋丽华,等.奶牛乳房炎大肠杆菌的血清型及耐药性调查研究[J].中国兽医杂志,2006,42(12):19-20.
[4]Santos J E P,Cerri R L A,Ballou M A,et a1.Effect of timing of first clinical mastitis occurrence on lactation and reproductive performance of Holstein dairy cows[J].Animal Reproduction Science,2004,80(122):31-45.
[5]Mathieu Brochet,Elisabeth Couve,Philippe Glaser,et al.Integrative conjugative elements and related elements are major contributors to the genome diversity of Streptococcus agalactiae[J].Journal of bacteriology,2008,10:6913-6917.
[6]Immuaculada Margarit,Cira Daniela Rinaudo,Cesira L.Galeotti,et al.Preveting bacterial infections with pilus-based vaccine:the group B Streptococcus paradigm[J].J Infect Dis,2009,199(1):108-115.
[7]Margarit I,Rinaudo CD,Galeotti C,et a1.Preventing bacterial infections with pilus-based vaccines:the group Bstreptococcus paradigm[J]..The Journal of Infectious Diseases 2009 Jan 1;199(1):108-15
[8]布日额,任晓峰,吴金花,等.牛乳腺炎无乳链球菌表面蛋白pgk抗原优势区的原核表达及其间接ELISA检测方法的建立[J].中国预防兽医学报,2013,35(4);312-6
[9]张海宝,布日额,王学理,等.牛乳腺炎无乳链球菌pgk基因的克隆与序列分析[J].中国病原微生物学杂志,2011,6(2);90-2
[10]郎景民,布日额,吴金花,等.牛乳腺炎无乳链球菌内蒙古分离株sip基因抗原表位序列的高效表达及其抗原性鉴定[J].中国病原微生物学杂志,2010,5(11);805-9.
[11]Telford JL,Barocchi MA,Margarit I,et al.Pili in Gram-positive pathogens[J].Nat Rev Microbiol,2006,4(7):509-19.
[12]Brintoncc,Bryan J,Dillon J,et al.Uses of pili in gonorrhea control;role of bacterial pili in disease,purification and properties of gonococcal pili,and progress in the development of a gonococcal pilus vaccine for gonorrhea.In Brooks G,Gotschlich EC,Holmes KK,Sawyer S and Young F(ed.)Immunobiology of Neisseria gonorrhoeae[M].American Society for Microbiology,1978.155-78.
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