肺炎支原体双向电泳条件的探索  

Optimization of two-dimensional polyacrylamide gel electrophoresis techniques for mycoplasma pneumoniae

作　　者：胡璇[1] 苑鑫[1] 柏长青[1] 牛文凯[1] 袁静[2] 冯羽中 李璞媛[1] 刘慧莹[1] HU Xuan YUAN Xin BAI Chang-qing et al(Department of Respiratory and Critical Care Medicine, the Affiliated Hospital of the Academy of Military Medical Science, Beijing 100071, China Institute of Disease Control and Prevention ,Academy of Military Medical Sciences ,Beijing 100071 ,China)

机构地区：[1]军事医学科学院附属医院呼吸与危重症医学科,北京100071 [2]军事医学科学院疾病预防控制所,北京100071

出　　处：《中国实验诊断学》2016年第12期2006-2008,共3页Chinese Journal of Laboratory Diagnosis

基　　金：首都特色基金(Z141107002514182);国家自然科学基金(81400009)

摘　　要：目的建立适用于肺炎支原体(mycoplasma pneumoniae,MP)的双向电泳技术。方法对双向电泳实验中的三个关键因素:全蛋白提取量、固相PH梯度胶条范围、蛋白上样量进行摸索,比较和分析标准株FH电泳图谱,利用液相色谱串联质谱技术(LC-MS/MS)鉴定部分蛋白斑点。结果采用菌液离心结合超声破碎的方法提取全蛋白,上样量为2mg,固相PH梯度胶条选取PH4-7,可以得到蛋白点分布均匀的双向电泳图谱,蛋白点数为60±2。利用液相色谱串联质谱(LC-MS/MS)技术成功获得了分子伴侣蛋白(Chaperone protein DnaK),鉴定成功率为100%。结论成功建立了适用于肺炎支原体蛋白质组分析的双向电泳技术条件,为肺炎支原体蛋白质组学的研究提供一个可靠的技术平台。Objective To develop a scheme of two-dimensional polyacrylamide gel electrophoresis（2-DE）for myco-plasma pneumoniae（MP）.Methods The main factors of 2-DE technique,including protein precipitation,PH gradient gel strip range and protein sample size,were studied in this research.The standard FH electrophoresis map was com-pared and analyzed,and then some protein spots were identified by liquid chromatography tandem mass spectrometry （LC-MS/MS）.Results The proteins spots of FH were well distributed under the condition of whole protein extracted by centrifugal and ultrasonic fragmentation,the 2mg protein sample size,and PH 4-7.60±2 protein spots were detected in the maps.By using LC-MS/MS,protein identification results were molecular chaperone protein DnaK.Identification rate was 100%.Conclusion The 2-DE method for proteomics analysis of MP was successfully established which could be effectively used in the MP proteomics investigation.

关 键 词：肺炎支原体 蛋白质组学 双向电泳技术 

分 类 号：R375.2[医药卫生—病原生物学]