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。
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 PAl03. The mutations in amino acid sequence are: Asn-364 to Ser; Ser-506 to Tip and Ser-515 to Gly. Conclusion: The mutations have a little influence on the enzymatic activity and cytotoxicity, so the gene PF40 of Pseudomonas aeruginosa ATCC27853 will be usefull in the research of molecular targeted protein.
出处
《中国农学通报》
CSCD
2006年第8期30-32,共3页
Chinese Agricultural Science Bulletin
基金
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)