摘要
目的提纯甲型副伤寒沙门菌鞭毛蛋白并鉴定。方法选用甲型副伤寒沙门菌,经自制的液体培养基扩大培养后,酸裂解法初步分离出鞭毛蛋白,用AKTAExplorer蛋白纯化系统除盐,弱阴离子交换层析纯化。纯化的蛋白用SDS-PAGE、Westernblot和磷钨酸负染扫描电子显微镜(scanningelectronmicrocopy,SEM)观察并摄片鉴定。考马斯亮蓝法测定所获得鞭毛蛋白的产量。结果SDS-PAGE提示纯化的鞭毛蛋白为一条相对分子质量(Mr)为52×103蛋白带;免疫印迹试验亦提示条带在Mr52×103处;SEM观察发现该鞭毛蛋白呈丝状;三者均证实该蛋白为同一均质蛋白。蛋白定量测得每克湿重的细菌可提取(4.8±0.5)mg鞭毛蛋白。结论经酸裂解法可获得高产量的鞭毛蛋白,且易于纯化和鉴定。
Objective To abstract and identify the flagellin from Salmonella paratyphl A. Methods Salmonella paratyphl A was obtained by centrifuging. Then, the flagellin was isolated by acid-splitting, and purified by AKTA Explorer protein purified system. The purified flagellin was identified by SDS-PAGE, Western blot, phosphotungstic acid negative staining and scanning electron microscopy. Results SDS-PAGE and Western blot showed that the purified flagellin protein strip was located at Mr 52× 10^3 region; the protein appeared filaceous in scanning electron microscopy, indicating that the purified flagellin was a homogeneous protein. One gram of wet Salmonella paratyphi A produced （4.8 ± 0.5） mg of freeze-dried flagellin. Conclusion The fruitful flagellar protein from Salmonella paratyphl A can be obtained by acid-splitting, which can be purified and identified easily.
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2005年第7期599-603,共5页
Chinese Journal of Microbiology and Immunology
基金
国家自然科学基金资助项目(30370631)
作者简介
通讯作者：徐剑铖，Email：xu822036@sohu．com，电话：023-68755005
