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介绍一种单孢直接挑取获得单孢菌株的方法 总被引：6，自引：3，他引：6
单孢分离是植物病原真菌研究中的一项基础技术,常用的有琼胶平板稀释纯化法、琼胶平板表面单孢挑取法、微块单孢分离法、印痕移取单孢分离法等,各有利弊,但均需预先制备琼胶平板。为简化单孢分离过程,参照国内外使用的方法,并根据我们的实验条件, 相似文献
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单孢分离是植物病原真菌菌种纯化过程中一项不可缺少的技术,常见的有采用琼胶平板稀释纯化法或琼胶平板表面单孢子挑取法,近车又有人提出微块单孢子分离法。平板稀释纯化的可靠性较差,很难达到单孢子分离的目的;单孢子挑取可靠性虽高于前者,但不易掌握;微块法优于前两种方法,但获得单孢子的机率低,仅有10—20％。我们在菌种纯化的试验研究中,综合了目前较常用的几种方法,设计成印痕移取单孢子分离法。 一、操作器材的制备 (一)印痕器:由塑料套筒、胶塞、印痕筒3部分组成。 1.塑料套筒:根据所用显微镜低倍镜的外径大小,选择(可用试剂瓶的塑料内塞代 相似文献
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为从河西走廊石羊河流域地区盐碱土中分离筛选对动物病原菌具有抑菌活性的放线菌菌株。采用稀释平板涂布法、琼脂块法和发酵液滤纸片法,对盐碱土样中分离纯化得到的298株放线菌进行抗菌活性筛选,并对具有较高抑菌效果的放线菌菌株进行形态特征、生理生化特性及16S rRNA基因序列系统发育分析鉴定。研究表明:从盐碱土中分离的298株放线菌中,占35.2%的放线菌具有不同程度的抑菌效果;通过初筛和复筛得到11株抑菌效果较好的放线菌菌株,结合形态特征、生理生化特性及16S rRNA基因序列系统发育分析初步鉴定,其均为链霉菌属(Streptomyces)。河西走廊石羊河流域盐碱土中存在大量具有抗动物病原菌的放线菌,这为进一步发掘该地区放线菌资源提供了基础数据。 相似文献
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茶树叶面真菌与内生真菌群落结构的比较 总被引：2，自引：0，他引：2
采用常规真菌分离法对健康茶树叶片叶面真菌和内生真菌进行分离,比较了叶面真菌和内生真菌的种群和数量差异。结果表明：从茶树品种黄旦（HD）共分离得到叶面真菌26个形态型,内生真菌18个形态型;从铁观音（TGY）共分离获得叶面真菌16个形态型,内生真菌15个形态型：分离所得真菌大多数为半知菌和子囊菌,少数为担子菌。采用稀释平板法和叶片组织黏附法两种不同的真菌分离方法,获得叶面真菌优势种的种类基本一致,但各种菌的数量差异明显;对黄旦和铁观音两品种比较,采用叶片组织黏附法分离所得叶面真菌的数量没有差异,但采用稀释平板法所获不同真菌的数量差异明显。真菌分离结果表明在茶树叶片表面和内部组织中生长有显著不同的真菌群落。 相似文献
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为筛选出对肿节风炭疽病菌Colletotrichum dematium具有拮抗作用的细菌,开发新型生物制剂,本试验分别采用组织分离法和稀释平板法,从广西药用植物园科研基地健康肿节风植株的不同组织和根系土壤分离、纯化获得49株细菌。平板对峙试验结果表明,来自土壤的zjt-9、zjt-4,来自根系的zgs21及叶片的zyy13对肿节风炭疽病菌具有较强的拮抗作用,其中作用最强的是zjt-9。抗菌谱测定结果表明, zjt-9、zjt-4和zyy13对供试的18种病原真菌均有明显的拮抗效果,拮抗作用最强的是zjt-9,平均抑菌宽度最高达19.10 mm;经显微镜观察,拮抗细菌可造成肿节风炭疽病病原菌菌丝膨大、畸形、断裂。通过形态学特性和16S rDNA鉴定,zjt-9菌株为甲基营养型芽胞杆菌Bacillus methylotrophicus。 相似文献
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通过对不同作物根际土壤的处理，分离纯化出９２株细菌，从平板对峙培养和菌液对致病镰刀菌的抗生作用实验结果中，进一步筛选出１２株对三种致病镰刀菌生长有抑制作用的拮抗菌。 相似文献
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单孢分离是植物病理学的基本技术 ,它可纯化病原菌 ,是进行病原鉴定、生理生化、病菌遗传变异等方面研究的基础。单孢分离的方法很多 ,常采用ＰＤＡ平板划线法。但这种方法不太适用于马铃薯晚疫病菌 ,因为 :(1)市售琼脂杂质较多 ,经高压灭菌后仍可保留许多不规则的小块 ,在显微 相似文献
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为了分析草莓镶脉病毒（Strawberry vein banding virus,SVBV）ORF Ⅵ基因的功能,采用原核表达技术获得该基因表达的重组蛋白并制备其抗血清。利用PCR技术扩增得到SVBV中国分离物的ORF Ⅵ基因,将此基因克隆到原核表达载体pET-SUMO上,获得重组质粒pET-SUMO-ORF Ⅵ。转化大肠杆菌BL21（DE3）后,经IPTG诱导与Ni2+-NTA亲和柱纯化,获得分子质量约为90 kD的重组蛋白。以纯化的重组蛋白为抗原免疫家兔制备抗血清,采用间接ELISA和Western blot方法测定抗血清的效价、反应灵敏度以及ORF Ⅵ基因在本氏烟中的瞬时表达。结果显示,间接ELISA法测定抗血清对重组蛋白的效价达1：256 000,Western blot法能够检测到稀释64 000倍的重组蛋白。利用稀释2 000倍的抗血清,仍能够检测出SVBV中国分离物和美国分离物ORF Ⅵ基因在本氏烟中瞬时表达的蛋白。 相似文献
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ABSTRACT A new technique, the spiral gradient dilution method, was evaluated for determining 50% effective concentrations (EC(50) values) of fungicides for the inhibition of mycelial growth and conidial germination of various fungi. In this method, an agar medium is plated with a fungicide solution by means of a spiral plater, which applies the fungicide in a 2.5-log dilution in a continuous radial concentration gradient. Fungal inoculum is then placed along radial lines across the gradient. After incubation of the plates, distinct growth shapes were observed in different fungus-fungicide interactions. Mycelial growth responses to increasing fungicide concentrations ranged from abrupt to gradual transitions. Conidial germination responses were similar; in addition, distinct zones of confluent growth, nonconfluent growth, and outlier colonies were also identified, depending on the fungus-fungicide interaction. The fungicide concentration at the radial distance at which 50% reduction of growth or spore germination occurred, compared with growth or germination on unamended media, was calculated by a computer program. EC(50) values were obtained for mycelial growth in 22 fungus-fungicide interactions and for conidial germination in five interactions. The fungi evaluated were members of the Zygomycota, Ascomycota, Basidiomycota, and Deuteromycota. Nine fungicides, belonging to six different chemical classes, were tested. EC(50) values were compared with those obtained by the traditional agar dilution method. In linear regression analyses of the two methods, the models were highly significant (P < 0.01), and coefficients of determination (r(2)) were 0.92 for the mycelial growth assays and 0.94 for the conidial germination assays. Regression slopes were not significantly different from 1 (P > 0.05) with optimal program settings in the software. Estimated bias, coefficients of variation, and actual confidence intervals for the regression slope were 13.5%, 6.5%, and 1.14 +/- 0.14 for the mycelial growth assays and 7.5%, 14.5%, and 1.08 +/- 0.37 for the conidial germination assays. These analyses indicate that the spiral gradient dilution method is accurate and precise compared with the agar dilution method for calculating EC(50) values of fungicides in continuous growth responses to fungicide concentration gradients. 相似文献
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三种三唑类杀菌剂对映体生物活性的研究 总被引：3，自引：5，他引：3
采用高效液相色谱法(HPLC)拆分了戊唑醇(tebuconazole)、粉唑醇(flutriafol)和己唑醇(hexaconazole)3种杀菌剂的外消旋体,并且制备了对映体的纯品。以花生褐斑病菌Cercospora arachidicola、番茄早疫病菌Alternaria solani、油菜菌核病菌Sclerotinia sclerotiorum、苹果轮斑病菌Alternaria mali、甜菜褐斑病菌Cercospora beticola等5种植物病原菌为供试菌,采用琼脂稀释法测定了3种杀菌剂的外消旋体及对映体的生物活性,比较了对映体之间抗菌活性的差异。结果表明:(-)-戊唑醇的抗菌活性优于(+)-戊唑醇;(+)-粉唑醇的抗菌活性优于(-)-粉唑醇;(-)-己唑醇的抗菌活性优于(+)-己唑醇。 相似文献
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分离土壤棉花黄萎病菌选择性培养基的筛选 总被引：8，自引：1，他引：8
采用土壤检测稀释法、水筛法和不同土样接种量,对5种土壤浸提液改良培养基进行了比较研究。结果表明,土壤水筛法对土壤棉花黄萎病菌的检测效果优于稀释法,其菌落检测数增加4~10倍。培养基每皿0.25 g土样接种量的检测效果高于0.5 g和1g。5种测试培养基中,培养基D对分离土壤棉花黄萎病菌的选择性表现最好,其土壤接种体检测回收率达63.5%~83.3%.培养基D的组份为:土壤浸提液25 ml,KH2PO4 1.5 g,K2HPO4 4 g,尿酸钠0.2 g,半乳糖醛酸钠(果胶)1g,山梨糖1g,Dox盐2 ml,Tergitol NP-101ml,PCNB0.1g,琼脂17g,蒸馏水1000 ml,pH6.4~6.7。每1000ml融化培养基倒皿前加抗菌素液50 ml (含氯霉素0.05 g,链霉素0.05 g,青霉素-G盐0.05 g)。 相似文献
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The antifungal activity of 61 N-phenylsuccinimides and 16 N-phenyl-1,2-dimethylcyclopropanedicarboximides having various benzene ring substituents was determined against Botrytis cinerea by the agar medium dilution method. The structure-activity relationships were analyzed using such physicochemical substituent parameters as hydrophobic π, electronic σ0, steric E8, and HB (hydrogen bonding) values with the multiple regression technique. The π values were derived from log P (octanol-water partition coefficient) values for the N-monosubstituted-phenylsuccinimide system. The hydrophobic effect is significant only for m-substitutents. The stronger the electron withdrawal and the smaller the steric dimensions of the ring substituents, the greater is the activity. When substituents are hydrogen bond acceptors, the effect is to lower the activity. These features are almost identical between two series of compounds. 相似文献
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Junya Takahashi Osamu Kirino Chiyozo Takayama Shigeko Nakamura Hiroshi Noguchi Toshiro Kato Katsuzo Kamoshita 《Pesticide biochemistry and physiology》1988,30(3)
For 69 methyl N-phenylcarbamates having various benzene ring substituents, the fungicidal activity was determined against Botrytis cinerea resistant to benzimidazole fungicides by the agar medium dilution method. The structure-activity relationships were analyzed quantitatively using such physicochemical substituent parameters as hydrophobic π, steric B5, and HB (hydrogen bonding) with the Hansch-Fujita method (C. Hansch and T. Fujita, J. Amer. Chem. Soc.86, 1616 1964). The hydrophobicity of substituents was favorable to the acitivity. The effect was position-specific, the importance being in the order of ortho meta > para. The activity was related parabolically to the maximum width of one of the m-substituents having the larger dimension. The hydrogen acceptability of p-substituents enhanced the activity. The preventive activity of compounds against gray mold of cucumber caused by the resistant B. cinerea, which was determined by the foliar application in pot tests, was dependent on the magnitude of the fungicidal activity and hydrophobicity of the compounds as analyzed by the adaptive least-squares method. 相似文献
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小麦茎基腐病生防菌株YB-161的分离鉴定及防效测定 总被引：4，自引：0，他引：4
为筛选防治小麦茎基腐病的高效生防菌株,采用稀释涂布法从小麦茎基腐病病土中分离细菌,以假禾谷镰孢Fusarium pseudograminearum为靶标菌,通过平板对峙培养法和室内盆栽试验筛选优良菌株并测定其田间防治效果,分析优良菌株发酵滤液对假禾谷镰孢的抑菌能力,并结合形态特征、Biolog微生物自动鉴定系统和gyrA序列分析对其进行种类鉴定,以及测定该菌株的抑菌谱。结果表明,10份供试土样中共分离到98株细菌,对假禾谷镰孢的菌丝生长抑制率超过40.00%的菌株有14株,其中菌株YB-161对假禾谷镰孢的菌丝生长抑制率最高,达63.50%。室内盆栽试验结果显示,菌株YB-161处理小麦的病情指数最低,为31.40,对小麦茎基腐病的防治效果最好,为68.15%,其对小麦也表现出较好的促生作用。田间调查结果显示,菌株YB-161菌液拌种处理对小麦茎基腐病的田间防治效果高于52.35%,并对小麦具有一定的增产作用。菌株YB-161的发酵滤液5倍稀释液和10倍稀释液对假禾谷镰孢的菌落抑制率分别达66.67%和44.33%,该菌株被鉴定为解淀粉芽胞杆菌Bacillus amyloliquefaciens。此外,菌株YB-161对辣椒炭疽病菌Colletotrichum capisci、烟草疫霉病菌Phytophthora parasitica等5种植物病原真菌均有较好的拮抗作用。表明生防菌株YB-161对绿色防控小麦茎基腐病具有良好的开发利用潜能。 相似文献
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J. W. L. VAN VUURDE 《EPPO Bulletin》1987,17(1):139-148
Antibodies coated onto a suitable solid phase, e. g. polystyrene beads, rods for inoculating agar plates, or petri dishes, enable selective trapping of homologous bacteria and of immunologically related bacteria by immunoaffinity. After washing away unbound organisms, the bound organisms are desorbed and plated on a suitable medium. Selective immunoisolation has been demonstrated for various phytopathogenic bacteria including Corynebacterium michiganense ssp. michiganense, Erwinia chrysanthemi, E. carotovora ssp. atroseptica, Xanthomonas campestris pv. begoniae and Pseudomonas syringae pv. phaseolicola. Three serological methods are described for additional rapid immunoidentification of colonies directly on agar plates and for screening for cross-reacting microorganisms: (a) direct immunodiffusion on dilution plates, (b) agar mixed-antibody assay, (c) fluorescent antibody colony staining. Schemes are presented for increasing the reliability and sensitivity of sample screening for quality indexing of plant material, and for efficient screening and isolation of possible cross-reacting microorganisms to enable production of more specific antisera. 相似文献
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菌株CQ是一株分离自云南省保山市昌宁县烟草根部的对番茄灰霉病具有较好防治效果的拮抗细菌。通过对该菌株进行形态特征观察、16S rDNA序列分析鉴定,确定菌株CQ为枯草芽孢杆菌Bacillus subtilis。拮抗试验结果表明,菌株CQ能够显著抑制番茄灰霉菌的菌丝生长,使菌丝出现异常分枝和囊泡状畸形。1‰ CQ发酵滤液即可显著抑制番茄灰霉菌孢子的形成。室内生测结果表明,相比对照组,48 h无菌发酵滤液原液在离体叶片上对番茄灰霉病的防效可达100%,而其10、100倍稀释滤液则使发病率分别降低28.5%和21.3%,病情指数分别降低44.01和37.86。 相似文献
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番茄早疫病菌拮抗放线菌10-4的鉴定 总被引：4，自引：2，他引：2
为明确放线菌10-4菌株在植物病害生物防治中的应用潜力,采用琼脂块法、菌丝生长速率法和温室盆栽试验测定其抑菌作用,并通过多相分类法研究其分类地位。结果表明:菌株10-4对20种植物病原真菌均有抑制作用,其中对小麦根腐病菌、稻瘟病菌、番茄早疫病菌、立枯丝核病菌、小麦全蚀病菌、苹果炭疽病菌和玉米大斑病菌的抑制作用较强,生长抑制率均达到90%以上;菌株10-4的代谢产物对植物病原真菌菌丝有致畸作用;其发酵原液和5倍稀释液在活体植株上对番茄早疫病具有良好的生防作用,防治效果分别为77.2%和70.9%。根据形态特征、培养性状、生理生化特性、细胞壁化学组分及16SrDNA序列分析结果,将菌株10-4初步鉴定为腐生链霉菌Strepto-myces saprophyticus。 相似文献