生物技术通讯
生物技術通訊
생물기술통신
LETTERS IN BIOTECHNOLOGY
2013年
5期
698-701
,共4页
胡桂萍%贾宪波%刘波%尤民生
鬍桂萍%賈憲波%劉波%尤民生
호계평%가헌파%류파%우민생
芽胞杆菌%蛋白提取%破碎方法
芽胞桿菌%蛋白提取%破碎方法
아포간균%단백제취%파쇄방법
Bacillus spp.%protein extraction%disruption
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。
目的：：比較不同破碎方法下3種芽胞桿菌菌體蛋白的提取效果。方法：以菊酯類農藥降解菌巨大芽胞桿菌、毬形芽胞桿菌、彎麯芽胞桿菌為研究對象，以裂解液為提取介質，分彆採用超聲波破碎法、水煮法和機械破碎法提取芽胞桿菌蛋白，併通過革蘭染色電鏡觀察菌株破碎程度，用SDS-PAGE和Bradford法測定蛋白濃度。結果：革蘭染色電鏡觀察髮現玻璃珠機械破碎法對3種芽孢桿菌的細胞壁破碎程度最明顯，其次為超聲波破碎法，水煮法破碎效果不明顯。SDS-PAGE和Bradford法測定結果錶明，巨大芽胞桿菌、毬形芽胞桿菌、彎麯芽胞桿菌經玻璃珠機械破碎後，提取的蛋白圖譜條帶清晰，豐度高，重複性好，含量分彆為20.247、19.902和18.893 mg/mL；經超聲波破碎提取的蛋白圖譜條帶較清晰，豐度一般，重複性不好，含量分彆為10.572、9.438和10.424 mg/mL；經沸水浴破碎提取的蛋白圖譜條帶模糊，豐度低，重複性差，含量分彆為1.366、1.119和1.136 mg/mL。結論：玻璃珠機械破碎法是破碎3種芽胞桿菌的最優方法，破碎後提取的蛋白含量高，條帶清晰，重複性好。
목적：：비교불동파쇄방법하3충아포간균균체단백적제취효과。방법：이국지류농약강해균거대아포간균、구형아포간균、만곡아포간균위연구대상，이렬해액위제취개질，분별채용초성파파쇄법、수자법화궤계파쇄법제취아포간균단백，병통과혁란염색전경관찰균주파쇄정도，용SDS-PAGE화Bradford법측정단백농도。결과：혁란염색전경관찰발현파리주궤계파쇄법대3충아포간균적세포벽파쇄정도최명현，기차위초성파파쇄법，수자법파쇄효과불명현。SDS-PAGE화Bradford법측정결과표명，거대아포간균、구형아포간균、만곡아포간균경파리주궤계파쇄후，제취적단백도보조대청석，봉도고，중복성호，함량분별위20.247、19.902화18.893 mg/mL；경초성파파쇄제취적단백도보조대교청석，봉도일반，중복성불호，함량분별위10.572、9.438화10.424 mg/mL；경비수욕파쇄제취적단백도보조대모호，봉도저，중복성차，함량분별위1.366、1.119화1.136 mg/mL。결론：파리주궤계파쇄법시파쇄3충아포간균적최우방법，파쇄후제취적단백함량고，조대청석，중복성호。
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.
