【作者】 孙长坡；

【导师】 刘训理；

【作者基本信息】 山东农业大学 ， 特种经济动物饲养， 2003， 硕士

【摘要】 从泰山林地土壤中分离获得一株多粘类芽孢杆菌（Paenibacillus polymyxa），该菌株对板栗干腐病菌（Diplodia maydis）、茶云纹叶枯病菌（Colletorot richam camelliae）、茶轮斑病菌（Physatospora piricola Nose.）、棉花立枯病菌（Rhizoctomia sola）玉米圆斑病菌（Bipolaris zeicola shoemaker）、辣椒疫病菌（Phyto phthora Casici Leon）、黄瓜枯萎病菌（Fusarium vasin fectum）、家蚕白僵菌（Beauveria bassiana）、苏云金芽孢杆菌（Bacillus thuringiensis var.kurstaki）、白假丝酵母（Candida albicans）和大肠杆菌（E.coli k12）等都有强烈拮抗作用。通过对该菌株进行形态观察、生理生化实验、G+Cmol%测定和16SrRNA碱基序列测定，鉴定该菌株为类芽孢杆菌属中的一株新的多粘类芽孢杆菌，定名为Paenibacillus polymyxa Cp_S316，编码其16SrRNA的基因为一新基因，已经提交Gene Bank注册，注册号为AY292989（2003年6月4日）。对该菌株发酵条件的研究表明：该菌株用马铃薯葡萄糖液体培养基发酵培养， 28℃，200rpm摇瓶中振荡培养72h可获得高活性的发酵产物，用苏云金芽孢杆菌HD-1做指示菌，将发酵液稀释40倍生测仍可形成明显的抑菌圈。以8000rpm对发酵液离心收集菌体。用蒸馏水对菌体稀释；加入适量吸附树脂在150rpm、28℃下振荡吸附4h， 80%的丙酮解吸，过滤解吸液得到活性物质的澄清溶液，旋转蒸发仪旋转蒸发去处丙酮，经SephadexG-75分子筛层析得单一活性峰 ，收集峰值部分样品液经冷冻干燥得到淡褐色粉末，该活性物质用丙酮充分洗涤、甲醇-乙醚重结晶获得略带微黄的白色粉末，该活性物质50μg/ml仍可对苏云金芽孢杆菌HD-1产生明显的抑制作用。该菌株产生的活性物质耐高温，121℃加热25min活性基本不丧失；对丙酮、乙醚、氯仿等有机溶剂不敏感。在酸性条件下，活性基本不变，但在强碱条件下没有活性（调回PH值可恢复活性）。更多还原

【Abstract】 A bacterium named Paneibacillus polymyxa is screened from soil sample of Mountain Tai. It secrets a large amounts of active substance which can inhibit against several plant and animal diseases，such as Candiada albicans 、Aspergillus flavus Link and Nomuraea rileyi Farlow .The determination show that it is a strain of paenibacillus polymyxa through measuring its G+Cmol% and the sequences of bases of 16SrRNA. The cultural conditions such as temperature, fermentation period and the compound of medium are studied. The result of test show that suitable factors for both bacterium to grow and active substance to produce are 28℃,200rpm and 72 hours. The bacterium is gotten through centrifuge with 8000rpm for 20min.Then the bacteria is diluted and colophony named S-8 is put into and used to absorbed active substance for 4 hours. The active substance is unshiped from colophony through acet. After the acet is vaporized, The active substance in water is gotten. And which is vaporized at low temperature .Then the crude active substance is purified by column chromatography on sephadex G-75.After a series of purifications again ,we could get some white powder at last .Though the active substance is diluted to50μg/ml, The activity is still checkeded-up through Phyto phtnora casicileon.The purified active substance is insensitive to heat, resistant to chloroform、ethanol and the orhers.In addition, the active substance is sensitive to high pH(10~14),but it is not sensitive to low pH(1~5).Furthermre, when the pH is made to low again , The activity of it’s comes back.更多还原