| 绿脓杆菌ATCC27853外毒素PE40基因的扩增与测序 |
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| 作者姓名： | 顾小龙 秦建华 张宁 郭莉 |
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| 作者单位： | 河北农业大学动物科技学院 河北保定071001(顾小龙，秦建华，郭莉)，河北省畜牧兽医研究所 河北保定071001(张宁) |
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| 基金项目： | 河北省自然科学基金资助项目“堆型艾美耳球虫特异性单抗-PE40重组毒素的构建”(303226) |
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| 摘 要： | 目的扩增绿脓杆菌ATCC27853外毒素基因PE40,并对PCR产物进行测序鉴定,为获得用于构建分子导向药物的毒素弹头基因PE40奠定基础。方法采用PCR技术,以绿脓杆菌ATCC27853基因组DNA为模板,扩增PE40基因并测序。用DNASTAR软件将测得的序列与GenBank中的国际标准产毒株PA103的PE40基因序列进行比较。结果扩增出目的基因片断,长度为1231bp。测序表明,ATCC27853与PA103核苷酸同源性为98%,产生了7个碱基的突变,突变碱基导致第364位的天冬酰胺变成丝氨酸,第506位的丝氨酸变成色氨酸,第515位的丝氨酸变成甘氨酸。但产生变化的3个氨基酸均不是文献报道中的重要活性位点。结论产生变化的3个氨基酸不会对PEA的酶活性及细胞毒性产生很大影响,绿脓杆菌ATCC27853的PE40基因可用作分子导向药物的弹头。
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| 关 键 词： | PEA(绿脓杆菌外毒素A) PE40(去掉细胞结合区Ia的外毒素基因) PCR(聚合酶链式反应) |
Amplification and Sequencing of Exotoxin A Structural gene PE40 of Pseudomonas Aeruginosa ATCC27853 |
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| Authors: | Gu Xiaolong Qin Jianhu Zhang Ning Guo Li |
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| Institution: | 1,College of Aruimal Science and Technology, Hebei Agricultural Uriversity, Baoding 071001, 2Dairy Farm of Hangu, Tianjin 300400, 3Hebei Institute of Husbandry and Vetednary, Baoding 071001 |
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| Abstract: | Objective: To analyse nucleotide sequence of exotoxin A structural gene PE40 of Pseudomonas aeruginosa ATCC27853. Method: The gene PE40 was amplified from genome DNA of Pseudomonas aeruginosa ATCC2785 by PCR technique. The PCR product was purified, sequenced and compared with a standard strain PA103 from GenBank by DNASTAR software. Result: An objective gene fragment with the length 1231bp was obtained. The nucleotide sequence analysis showed that ATCC27853 shared 98% alignment with PA103. The mutations in amino acid sequence are: Asn-364 to Ser; Ser-506 to Trp and Ser-515 to Gly. Conclusion: The mutations have a little influence on the enzymatic activity and cytotoxicity, so the gene PE40 of Pseudomonas aeruginosa ATCC27853 will be usefull in the research of molecular targeted protein. |
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| Keywords: | PEA(Pseudomonas aeruginosa exotoxin A) PE40(exotoxin A structural gene devoid of its native celling binding domain Ia) PCR(Polymerase chain reaction) |
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