贝氏考克斯体毒力与毒力相关蛋白芯片的研制  

Preparation of protein microarray of virulence and virulence-related proteins of Coxiella burnetii

作　　者：王锡乐[1] 温博海[1] 陈梅玲[1] 王君[1] 孙长俭[1] 杨晓[1] 

机构地区：[1]军事医学科学院微生物流行病研究所病原微生物生物安全国家重点实验室,北京100071

出　　处：《军事医学科学院院刊》2008年第2期121-125,共5页Bulletin of the Academy of Military Medical Sciences

基　　金：国家自然科学基金资助(30670101);国家自然科学基金青年基金(30700744)

摘　　要：目的:构建贝氏考克斯体毒力与毒力相关蛋白芯片。方法:根据贝氏考克斯体全基因组序列,筛选出约160个编码毒力及毒力相关蛋白的基因,采用PCR扩增基因片段,将获得的基因片段与pET32a表达载体连接,然后将该重组质粒转化大肠杆菌并使目的基因表达,采用Ni-NTA亲和层析柱纯化表达的目的重组蛋白。将纯化的重组蛋白点在醛基化玻片上制备蛋白芯片。结果:由贝氏考克斯体104个重组蛋白构建毒力与毒力相关蛋白芯片,通过荧光扫描仪分析蛋白芯片与贝氏考克斯体感染的小鼠血清反应,发现10个重组蛋白与该血清反应为强阳性。结论:所制备的毒力与毒力相关蛋白芯片具有贝氏考克斯体特异性,可用于贝氏考克斯体感染患者血清的检测。Objective：To establish a protein microarray with virulence and vimlence-related proteins of Coxiella burnetii, Methods： According to the genome sequence of C. burnetii. 160 genes encoding vindence and virulence-related proteins were selected. The selected genes amplified by PCR were inserted into pET32a vector and expressed in Eschericbia coli. The recombinant proteins were purified by Ni-NTA affinity chromatography and used to prepare microarray, Results： The protein microarray was constructed with 104 recombinant proteins suceessfully expressed in E. coli. Ten of the recombi- nant proteins in microarray were found to be strongly recognized by sera from mice experimentally infected with C. burnetii in scanning analysis with a laser scanner. Conclusion： The protein microarray is specifie to C. burnetii and may be used for analysis of the sera from patients infected with C. burnetii.

关 键 词：贝氏考克斯体 重组蛋白 蛋白芯片 

分 类 号：R378[医药卫生—病原生物学]