芽胞杆菌菌体蛋白提取方法的比较

目的：比较不同破碎方法下3种芽胞杆菌菌体蛋白的提取效果。方法：以菊酯类农药降解菌巨大芽胞杆菌、球形芽胞杆菌、弯曲芽胞杆菌为研究对象，以裂解液为提取介质，分别采用超声波破碎法、水煮法和机械破碎法提取芽胞杆菌蛋白，并通过革兰染色电镜观察菌株破碎程度，用SDS-PAGE和Bradford法测定蛋白浓度。结果：革兰染色电镜观察发现玻璃珠机械破碎法对3种芽孢杆菌的细胞壁破碎程度最明显，其次为超声波破碎法，水煮法破碎效果不明显。SDS-PAGE和Bradford法测定结果表明，巨大芽胞杆菌、球形芽胞杆菌、弯曲芽胞杆菌经玻璃珠机械破碎后，提取的蛋白图谱条带清晰，丰度高，重复性好，含量分别为20.247、19.902和18.893 mg/mL；经超声波破碎提取的蛋白图谱条带较清晰，丰度一般，重复性不好，含量分别为10.572、9.438和10.424 mg/mL；经沸水浴破碎提取的蛋白图谱条带模糊，丰度低，重复性差，含量分别为1.366、1.119和1.136 mg/mL。结论：玻璃珠机械破碎法是破碎3种芽胞杆菌的最优方法，破碎后提取的蛋白含量高，条带清晰，重复性好。

Comparison of the Methods to Extract Proteins from Bacillus spp.

Objective： To compare the cell disruption methods for extracting proteins from Bacillus megaterium, B. sphaericus and B.flexus. Methods： Three treatments with boiling water digestion, ultrasonic disruption and glass-beads grounding were carried out and the lysis solution was used as extracting medium in the experiment. The strains＇ crushing degree was judged by scanning electron microscope after gram staining, and the proteins were analyzed by SDS-PAGE and Bradford＇s protein determining method. Results： The strains＇ cytoderm was crushed most severely by glass-beads gounding, and then by ultrasonic disruption, the worst by boiling water diges-tion. The SDS-PAGE analysis showed that the protein pattern with clear banding, high abundance and good repeat-ability by glass-beads-gounding, and the protein concentrations were 20.247, 19.902 and 18.893 mg/mL respective-ly. The pattern with general clear banding and abundance and bad repeatability by ultrasonic disruptin, and the concentration were 10.572, 9.438 and 10.424 mg/mL. The obscure banding, low abundance, worst repeatability and the little concentration with 1.366, 1.119 and 1.136 mg/mL respectively by boiling water digestive. Conclusion：The results with high contents, distinct strips and good repeat showed that method of mechanical crushing extrac-tion was the best to extract the protein from Bacillus species.