二土壤细菌的分离与纯化.ppt

实验二 土壤细菌的分离与纯化 一 　教学要求 通过从土壤中分离纯化细菌，初步掌握微生物的分离纯化方法和无菌操作技术。 二　实验原理 - 常用的分离纯化方法 microorganisms exist in nature as mixed populations. However, to study microorganisms in the laboratory we must have them in the form of a pure culture, that is, one in which all organisms are descendants of the same organism. 三 材料与器材 金黄色葡萄球菌(Staphylococcus)和普通变形菌(Proteus vulgaris)斜面菌种。 已灭菌的牛肉膏、高氏1号、土豆蔗糖培养各1瓶，49.5 mL 无菌水（带玻璃珠）1 瓶、4.5 mL无菌水6 管、80%乙酸、10%酚酸、95% 乙醇。 无菌培养皿12套、1 mL无菌移液管10 支、土壤样品、天平等。 五 注意事项 制备土壤稀释液时，要注意使土样均匀分散在稀释液中。 注意无菌操作。 六 思考题 试拟出从土壤中分离芽孢杆菌的主要实验步骤。 如何检验平板上某个单菌落是不是纯培养？ * * 单细胞挑取法 简易单细胞挑取法 micromanipulator 平板分离法 streak plate method The most common way of separating bacterial cells on the agar surface to obtain isolated colonies is the streak plate method. It provides a simple and rapid method of diluting the sample by mechanical means. As the loop is streaked across the agar surface, more and more bacteria are rubbed off until individual separated organisms are deposited on the agar. After incubation, the area at the beginning of the streak pattern will show confluent growth, while the area near the end of the pattern should show discrete colonies. pour plate method Another method of separating bacteria is the pour plate method. With the pour plate method, the bacteria are mixed with melted agar until evenly distributed and separated throughout the liquid. The melted agar is then poured into an empty plate and allowed to solidify. After incubation, discrete bacterial colonies can then be found growing both on the agar and in the agar. spread plate method The spin plate method involves diluting the bacterial sample in tubes of sterile water, saline, or broth. Small samples of the diluted bacteria are then pipetted onto the surface of agar plates. A sterile, bent-glass rod is then used to spread the bacteria evenly over the entire agar surface. 四 操作步骤 制备土壤稀释液 倾注平板 培养 　接种和分离工具 1．接种针 2.接种环 3.接种钩 4.5.玻璃涂棒 6.接种圈 7.接种锄 8.小解剖刀 挑菌落 iso