正交设计优化青蒿SRAP-PCR反应体系及引物筛选

为了建立青蒿的SRAP最佳扩增体系,并筛选出SRAP多态性引物,本研究以青蒿叶片DNA为模板,采用正交试验设计,以Mg^2+、dNTP Mix、Taq DNA聚合酶、引物和DNA模板5种因素5个水平,对青蒿SRAP反应体系进行研究。结果表明,青蒿SRAP-PCR最佳反应体系为:引物0.6μmol/L、Mg^2+2.0 mmol/L、模板DNA 5.1 ng、Taq DNA聚合酶2.0 U、dNTPs 0.25 mmol/L,总体积为25μL。各因素对扩增反应均有不同影响,其中引物浓度的影响最大,dNTPs的影响最小。运用该体系对不同种质资源的青蒿进行验证,证明该体系稳定可靠,并在30个引物组合中筛选出了25对扩增条带清晰,多态性丰富的引物组合。这一结论为今后利用SRAP标记技术进行青蒿分子遗传学研究提供了科学依据。     

捕食线虫性真菌的分离培养及分布规律

对自然界中的捕食线虫性真菌进行了分离和种属区分，并对分布规律进行了研究。从土壤、粪便等材料中获得了2类活性较强的捕食线虫性菌株：即套捕型(hooping)捕食线虫性真菌和粘捕型(sticking)捕食线虫性真菌。主要的代表种类有3种。这些捕食线虫性真菌在捕食性结构分生孢子形态及某些生物学特性上有一定差异。套捕型捕食线虫性真菌代表种为少孢节丛孢菌(Arthrobotrys oligospara)和梨形指环菌(Dactylaria pyriformis)，它们以菌环、菌网作为捕食性结构(器官)，以套捕方式杀灭线虫幼虫。梨形指环菌可产生多量厚垣孢子(chlamyalospore)。粘捕型捕食线虫性真菌代表种为纺锤隔指孢菌(Dactylella ellipsospora)。可形成菌结捕食线虫性结构，以粘捕的方式杀灭线虫幼虫。结果表明：捕食线虫性真菌在土壤和家畜粪便中的分离率是40．41％；此类菌适合于生存在阴暗、潮湿、富含腐植质的环境中。在温暖季节时，采用0．4g／L玉米粉琼脂培养基(CMA)易于对其进行分离培养。     

捕食线虫性真菌菌种保存技术的研究

本试验对捕食线虫性真菌菌种保存技术进行了研究,测定了在不同介质保存条件下,捕食线虫真菌分生孢子和菌丝的存活情况及其不是寄生性线虫的能力.结果表明:在设定的保存条件下,分别经过1周冷冻融化或2周反复冷冻融化,被保存的分生孢子接种于玉米粉琼脂培养基上能生长发育:在加入线虫第3期幼虫后,能生产捕食器,对虫体发挥捕食作用.     

胃肠道线虫的黏膜免疫

从机体抗胃肠道线虫的黏膜免疫、胃肠道黏膜的抗原加工与递呈、胃肠道黏膜抵御寄生虫的效应机制等几方面论述了胃肠道线虫的免疫机理和免疫学研究的进展，旨在进一步阐明用免疫介入方法防治胃肠道线虫的理论基础，以解决胃肠道线虫治疗过程中的药物残留以及寄生虫的抗药性问题。     

猪带绦虫六钩蚴cDNA文库的构建与筛选

在体外将猪带绦虫虫卵孵化为有活力的六钩蚴。采用商品化试剂盒抽提六钩蚴总 RNA、m RNA,反转录为双链c DNA,再加 Eco R Adapter。在去除小片段后 ,dsc DNA与 λgt11噬菌体 DNA连接 ,经蛋白包装 ,构建了猪带绦虫六钩蚴λgt11c DNA文库。该文库效价为 3.5× 10 9pfu/ m L ,重组 DNA片段大小为 0 .5～ 3.2 kb,平均为 1.6 kb,蓝白斑比 1∶ 9。用抗体探针对文库进行免疫学筛选 ,在消除非特异性反应的基础上筛选约 10 6 重组子 ,共得到 118株强阳性克隆 ;应用 PCR鉴定上述部分阳性克隆 ,均扩增到 0 .5 kb以上的片段。结果显示 ,构建的文库合格 ,含六钩蚴所有抗原基因 ,可用于六钩蚴 c DNA克隆的筛选 ;用免疫学与 PCR联合筛选 c DNA文库 ,可消除假阳性     

Rapid Screening for Aluminum Tolerance in Cereals by Use of the Chlorophyll Fluorescence Test

The effects of aluminum on the photosynthetic apparatus were examined in cereals grown in nutrient solutions (pH 4.5) at two Al levels (0 and 148 μM). The chlorophyll fluorescence kinetics results confirmed that the soft wheat ‘BHG’ cultivar has the potential for growth on acid soils while triticale cultivars ‘Niovi’ and ‘Dada’ appeared to be relatively tolerant. The percentage decrease in F_v/F_m of the less tolerant cultivars after Al-treatment indicated a decrease in the efficiency of the primary photochemistry of PS II, while the decrease in the ratio F_V/F_osuggested that exposure of the cultivars ‘Dio’ and ‘Appulo E’ to aluminum caused injury to the thylakoid structure. The percentage fluctuations of the ratio F_v/F_mwere shown to correlate very closely with the assessment of injury as evaluated by the relative top fresh weight. Measurements of chlorophyll fluorescence in vivo could be used to monitor injury caused by “Al-stress”, and thus they may serve as a rapid screening test for Al tolerance.     

Development of a semi-selective medium and an immunofluorescence colony-staining procedure for the detection of Clavibacter michiganensis subsp. sepedonicus in cattle manure slurry

Various compounds and basal media were tested for their suitability to create a semi-selective medium for isolation ofClavibacter michiganensis subsp.sepedonicus (Cms) from cattle manure slurry containing c. 10⁸ colony forming units (cfu) per ml.Plating efficiency of Cms in yeast glucose mineral medium (YGM) was 104% compared with yeast peptone glucose medium. Nalidixic acid, polymyxin B sulphate and the experimental disinfectant S-0208 inhibited colony growth of cattle slurry bacteria as compared with Cms in YGM. The optimal concentration of these inhibitors in combination was determined by modified agar diffusion tests and by pour plating in 24-well tissue culture plates. The semi-selective medium YGMI consisted of YGM supplemented with nalidixic acid (2 mg/l), polymyxin B sulphate (30 mg/l) and S-0208 (125 mg/l). Plating efficiency varied for Cms between 50.9 and 69.6%, for cattle slurry bacteria between 1.8 and 2.5% and for saprophytes from potato heel end extracts between 11.5 and 27.4%.Differentiation of Cms colonies from other colonies was based on their small and bluish colony morphology in pour plates and on immunofluorescence colony-staining (IFC). IFC of a pure culture of micro colonies of Cms in YGM was possible after one day incubation (colonies c. 5 cells). Green background fluorescence in the agar gels was prevented by addition of Tween 20 (0.1%) to the washing buffer and the use of 1% agar gels. IFC of macro colonies of Cms in YGMI, visible with 4x objective magnification, was possible after 4 days. The detection level of the target organism in artificially inoculated cattle slurry in YGMI based on colony morphology varied between 1.4×10³ and 2.3×10⁴ cfu per ml of cattle slurry. Miniaturized plating combined with IFC, using wells in tissue culture plates (=16 mm), proved suitable for detection, but was c. 30 times les sensitive. The recovery of Cms was negatively correlated with the number of saprophytic colonies in the agar plates (R ²=0.74).     

Spring invasion of Heterodera schachtii in sugarbeet; a simulation study

A simulation model was developed for the spring invasion of the beet cyst nematode,Heterodera schachtii Schmidt, into sugarbeet roots, according to the state variable approach. This model describes the processes of egghatch, emergence of second stage larvae from cysts, migration to roots and penetration into roots quantitatively, using published data.In 1983 a field experiment was conducted to test this model.H. schachtii cysts were introduced at depths 6–29 cm in PVC-cylinders, buried in the soil. The rooting depth of sugarbeet seedlings, growing in these cylinders, was limited to 5 cm by 50 m mesh nylon gauze. Every 10 days the second stage larvae, which had penetrated into the roots of these seedlings were counted. After 50 days, about 40% of the eggs had hatched. More than 20% of the emerged larvae penetrated if the cysts had been buried undeeply, and only 4% if the cysts had been buried at 29 cm depth.The model predicted the course of penetration into the root during the first 40 days with reasonable accuracy (r²=0.79), but in the 5th period of 10 days the model made an overestimation of more than 100%. Egghatch after 50 days was correctly simulated. The differences in penetration into the root between the model and the experiment might result from an oversimplified simulation of the penetration success or the neglection of mortality of second stage larvae. Detailed experiments should be done to provide better parameters for these factors.Samenvatting Volgens de toestandsvariabele-benadering werd een simulatiemodel ontwikkeld van de voorjaarspenetratie van het bietecystenaaltje. Het model beschrijft aan de hand van literatuurgegevens het uikomen van de eieren, het verlaten van de cyst door de larven, de migratie naar en de penetratie in de wortel.In 1983 werd een veldproef uitgevoerd om het model te toetsen. Cysten vanH. schachtii werden op 5 dieptes tussen 6 en 29 cm ingegraven in PVC-cylinders, welke waren verzonken in de bodem. De bewortelingsdiepte van de suikerbiete-zaailingen die hierin groeiden werd beperkt tot 5 cm door nylon gaas van 50 m maaswijdte. Elke 10 dagen werden de larven geteld die in de wortels van deze plantjes waren gepenetreerd. Na 50 dagen was 40% van de eieren uitgekomen. Meer dan 20% van de gelokte larven penetreerden als de cysten ondiep waren ingegraven, en slechts 4% als de cysten op 29 cm diepte waren ingegraven.Gedurende de eerste 40 dagen werd het verloop van de penetratie in de wortel met redelijke nauwkeurigheid door het model voorspeld (r²=0.79). In de 5e periode van 10 dagen maakte het model echter een overschatting van meer dan 100%. Het uitkomen van de eieren werd correct gesimuleerd. De verschillen in penetratie tussen het model en de proef zouden het gevolg kunnen zijn van een oververeenvoudigde simulatie van het penetratiesucces of van het verwaarlozen van de mortaliteit van de migrerende larven. Betere gegevens hierover zullen moeten komen uit detailproeven.     

关于编制我国果树种质资源评价系统若干问题的商榷

资源评价是果树种质资源工作的中心环节。编制果树种资源评价系统是发展和提高我国果树种质资源工作水平的一项重要基础工作。“七五”期间,“果树种质资源主要性状鉴定评价”专题组编制１８种果树的《果树种质资源描述符》,于１９９０年在农业出版社出版。本文就其中其中若干问题提出商榷意见。     

草菇栽培原料发酵剂菌株的筛选及应用初探

从广西军区食用菌生产基地、广西大学微生物厂、广西大学牧场等地采集的样品中分离得到多株菌株。再从耐热、耐碱、产酶能力及营养物综合利用等方面考查各菌株，最终筛选得到理想的放线菌1株、细菌3株组成发酵剂菌株，并用于栽培原料发酵后栽培草菇。实验结果显示：加入该菌剂的料堆27h后温度可升至65℃，此时对照55℃，环境40℃，草菇生物学效率比对照提高了4.76％至7．56％。