用单克隆抗体作芜菁花叶病毒不同分离物外壳蛋白抗原结构的比较研究

用芜菁花叶病毒辽宁分离物（ＴｕＭＶ－ＬＮ）,山东分离物（ＴｕＭＶ－ＳＤ）和榨菜分离物（ＴｕＭＶ－ＺＣ）作抗原,分别免疫ＢＡＬＢ／ｃ小鼠,经脾细胞与ＳＰ２／Ｏ－Ａｇ１４骨髓细胞融合,共获得７个单克隆抗体分泌细胞株,为研究３个分离物的抗原差异,用胰蛋白酶消化ＴｕＭＶ的外壳蛋白（ＣＰ）。分别形成４、２和１条降解带,经ＳＤＳ－１５％ＰＡＧＥ后,转移至硝酸纤维薄膜上,用３个分离物的抗血清和７种单隆抗体分别做     

蚕豆萎蔫病毒纯化和病毒蛋白质及RNA组份分析

属蚕豆萎蔫病毒（ＢＢＷＶ）血清ＩＩ型的分离物Ｂ－９３４接种昆诺藜,采收的病叶先用含蔗糖的高浓度磷酸钾盐缓冲液,后用ＴｒｉｔｏｎＸ－１００处理,成功地提取了大量病毒粒子,提纯病毒得率为１３４ｍｇ／ｋｇ病叶,提纯病毒在电镜下可观察到大量球状病毒粒子,直径约２５ｎｍ,提纯的ＢＢＷＶ经甘油梯度超离心后在可观察到３个组份,分别称Ｔ,Ｂ和Ｍ组份,其中Ｔ为空壳蛋白,Ｂ和Ｍ为核蛋白,各组份经ＳＤＳ－ＰＡＧＥ测定,     

抗芜菁花叶病毒转基因甘蓝型油菜的研究

以子叶柄为材料,建立了甘蓝型油菜（ＢｒａｓｓｉｃａｎａｐｕｓＬ．）双低品种的再生体系。通过子叶柄与农杆菌（ＡｇｒｏｂａｃｔｅｒｉｕｍｔｕｍｅｆａｃｉｅｎｓＬＢＡ４４０４）共培养,将表达载体ｐＢＴｕ中芜菁花叶病毒外壳蛋白（ＴｕＭＶ－ＣＰ）基因以整合方式导入甘蓝型油菜,然后用卡那霉素进行筛选,获得了油菜再生植株。经ＰＣＲ特异性扩增、点杂交和Ｓｏｕｔｈｅｒｎ印迹分析,证明再生植株基因组ＤＮＡ中整合了ＴｕＭＶ－ＣＰ基因。攻毒实验表明,有ＴｕＭＶ－ＣＰ基因插入的工程油菜对ＴｕＭＶ均有不同程度的抗性。     

头端延髓腹外侧区血管加压素在刺激中脑dPAG区诱发防御性升压反应中的…

雄性Ｓｐｒａｇｕｅ－Ｄａｗｌｅｙ大鼠,用乌拉坦（７００ｍｇ／ｋｇ）和氯醛糖（３０ｍｇ／ｋｇ）腹腔麻醉。在双侧头端延髓腹外侧区（ｒＶＬＭ区）每侧微量注射血管加压素（ＡＶＰ）（１０ｐｍｏｌ／０．１μｌ）可引起平均动脉压（ＭＢＰ）升高,心率（ＨＲ）变化不明显,每侧微量注射ＡＶＰ的Ｖ１受体拮抗剂ｄ（ＣＨ２）５［Ｔｙｒ（Ｍｅ）＾２］ＡＶＰ（０．１ｎｍｏｌ／０．１μｌ）后ＭＢＰ和ＨＲ无明显变化。若预先在ｒＶＬ     

香蕉试管苗黄瓜花叶病毒检疫检验技术研究

建立了香蕉试管苗黄瓜花叶病毒ＤＡＣ－ＥＬＩＳＡ检测技术。应用０．１ｍｏｌ／Ｌ磷酸盐缓冲液（ｐＨ７．２,内含１％ＮａＤＩＥＣＡ）作为样品制备缓冲液,检测灵敏度可达到１／５１２０（稀释度）。两年来检测了１２８７个样品,香蕉试管苗生产繁殖材料的带毒率为２．８％。     

高效价甘薯羽状斑驳病毒抗血清的制备

用嫁接方法将甘薯羽状斑驳病毒（ＳＰＦＭＶ）接种到Ｉ．ｓｅｔｏｓａ上扩繁,以０．２ｍｏｌ／ＬｐＨ７．２ＰＢＫ缓冲液、垫层差速离心、蔗糖密度梯度离心提取纯化ＳＰＦＭＶ。纯化的ＳＰＦＭＶＯＤ２６０／２８０的比值为１．２５。将纯化的ＳＰＦＭＶ免疫家兔制备抗血清,在环状沉淀和微量沉淀试验中,用提纯病毒测定抗血清的效价均为１：４０９６;以ＳＰＦＭＶ－ＩｇＧ为第一抗体,应用Ｄｏｔ－ＥＬＩＳＡ对甘薯和Ｉ．ｓｅｌｏｓａ叶片中的ＳＰＦＭＶ分别作了测定。     

温州蜜柑萎缩病毒提纯与抗血清制作及其应用

采用洋酸浆（Ｐｈｙｓａｌｉｓｆｏｒｉｄａｎａ）繁殖温州蜜柑萎缩病毒（ＳＤＶ）,改进提纯方法,经５％－２５％,酒石酸钾密度梯度离心获得的提纯制剂,在２２０－３００ｎｍ下呈单一吸收峰曲线,峰在２５９ｎｍ,谷在２３７ｎｍ,ＯＤ２５９／ＯＤ２３７＝１．６０,ＯＤ２５９／ＯＤ２８０＝１．７３,病毒得率约为１３ｍｇ／ｋｇ病叶,电镜观察各视野布满了直径约２６ｎｍ的球状病毒颗粒,病毒粒子外壳蛋白有两个组分,分子量     

小麦黄花叶病毒RNA228kDa蛋白基因的表达及表达产物抗血清的制备

根据小麦黄花叶病毒（ Ｗ Ｙ Ｍ Ｖ） 核苷酸序列测定结果,将 Ｗ Ｙ Ｍ Ｖ Ｒ Ｎ Ａ２ 上的２８ ｋ Ｄａ 蛋白基因克隆到ｐ Ｅ Ｔ１１ａ 上,构建了原核表达载体ｐ Ｅ２８３９ 。 Ｓ Ｄ Ｓ Ｐ Ａ Ｇ Ｅ 分析表明,经 Ｉ Ｐ Ｔ Ｇ 诱导,２８ ｋ Ｄａ蛋白基因在大肠杆菌 Ｂ Ｌ２１（ Ｄ Ｅ３）ｐ Ｌｙｓ Ｓ 中得到高效表达。以含表达产物的凝胶为抗原,免疫家兔,首次制备了小麦黄花叶病毒 Ｒ Ｎ Ａ２ 蛋白特异性抗血清。     

兔迷走复合区内微量注射TRH对奥迪氏括约肌肌电的影响及传出途径分析

目的和方法：本工作旨在研究免迷走复合区（ＤＶＣ）内ＴＲＨ对奥迪氏括约肌（ＳＯ）肌电的调节作用及外周途径。结果：（１）ＤＶＣ内注射ＴＲＨ（０．８ｎｍｏｌ,１μｌ）后,慢波电位（ＳＷ）频率不变,但锋电位频率（ＦＳＰＳＯ）及幅度（ＡＳＰＳＯ）、锋电位发生率（ＩＳＰ）明显增加．（２）ＤＶＣ内分别注射不同剂量ＴＲＨ（０．１３,０．２５,０．５０,０．８０,１．３０ｎｍｏｌ,１μｌ）后,各剂量ＴＲＨ均能引起ＦＳＰＳＯ增加。随注射剂量的增加,ＳＯ反应强度和持续时间均增加,呈现明显的剂量反应关系。（３）ＤＶＣ内注射ＴＲＨ兴奋ＦＳＰＳＯ的效应可被静脉注射阿托品（０．２ｍｇ／ｋｇ）或迷走神经切断完全阻断,但不能被静脉注射酚妥拉明（１．５ｍｇ／ｋｇ）、心得安（１．５ｍｇ／ｋｇ）或脊髓切断术所阻断。结论：ＤＶＣ内ＴＲＨ可能对ＳＯ肌电有重要的调节作用,这种作用是通过迷走神经及外周Ｍ受体介导。     

侵染半夏的两种病毒的分离纯化和初步鉴定

用自然感染的半夏（Ｐｉｎｅｌｌｉａｔｅｒｎａｔａ）为材料,经粗提纯后检查到一种线状病毒和一种球状病毒,用两种方法对担提纯样品中的病毒粒子进行了分离纯化。１０％－７０％连续甘油梯度８００００ｇ离心１５０分钟获得两条病毒带,经紫外吸收测定均为强的核蛋白吸收峰,病毒粒子检查分别为球状和线状病毒粒子,线状病毒经浓缩收集为均一成份．与芋花叶病毒（Ｄａｓｈｅｅｍｍｏｓａｉｃｖｉｒｕｓ,ＤＭＶ）抗体有强的阳性反应。粗提纯样品经０．８％琼脂糖凝胶电脉分离为一条蛋白带,该条带回收后经紫外吸收测定为核蛋白吸收峰,电镜下检查为均一的球状病毒,以ＴＭＶ为对照、醋酸铀（ＵＡＣ）负染后测得该球状病毒（ｐｉｎｅｌｌｉａｓｐｈｅｒｉｃａｌｖｉｒｕｓ１．ＰｓＶ－１）的大小为３１．７ｎｍ;戊二醛固定后磷钨酸（ＰＴＡ）负染测得ＰｓＶ—１的大小为３４．０ｎｍ。各组分经ＳＤＳ－聚丙烯酸胺凝焦电泳分析测得线状病毒和球状病毒的外壳蛋白分子量分别为２０ＫＤ和２８ＫＤ。初步确定线状病毒为ＤＭＶ．球状病毒ＰｓＶ—１为侵梁天南星科半夏的一种新病毒。     

Disease intensity of some tomato viroses in Serbia

In this paper we present the data on the disease intensity of the tomato plants grown in glass and plastic-houses, and in the open field. The infection was caused by the following viruses: Tomato mosaic virus (ToMV), Tobacco mosaic virus (TMV), Tomato spotted wilt virus (TSWV), Alfalfa mosaic virus (AMV), Potato virus X (PVX), Potato virus Y (PVY), Tomato black ring virus (TBRV), Tomato ringspot virus (ToRSV), Tomato aspermy virus (TAV), and Cucumber mosaic virus (CMV). These viruses represented most frequent tomato pathogens in Serbia. According to the obtained results, it could be concluded that 92.94% of the tested tomato plants grown in glass and plastic-houses, and 89.82% grown in the open field were infected by one of the above viruses. Most of the plant samples were infected by two or more viruses. The most frequent viruses — tomato pathogens in Serbia were ToMV, PVY and TMV.     

Virus Resistance in Cereals: Sources of Resistance, Genetics and Breeding

In cereals, soil-borne viruses transmitted by the plasmodiophorid Polymyxa graminis (e.g., Barley mild mosaic virus , Barley yellow mosaic virus or Soil-borne cereal mosaic virus ), have increased in importance due to the increase of the acreage infested and because yield losses cannot be prevented by chemical measures. Due to global warming, it is also expected that insect transmitted viruses vectored by aphids (e.g., Barley yellow dwarf virus , Cereal yellow dwarf virus ), leafhoppers ( Wheat dwarf virus ) or mites (e.g., Wheat streak mosaic virus ), will become much more important even in cooler regions. The environmentally most sound and also most cost effective approach to prevent high yield losses caused by these viruses is breeding for resistance. Therefore, in contrast to other reviews on cereal viruses, this study briefly reviews present knowledge on cereal-infecting viruses and emphasizes especially the sources of resistance or tolerance to these viruses and their use in molecular breeding schemes.     

Identification of Peanut Green Mosaic Virus Strains in India*

During field surveys, three peanut green mosaic virus isolates differing in symptomatology on groundnut and a few other hosts were collected. Ultrathin sections of infected groundnut leaflets showed cytoplasmic inclusions with pin wheels and scrolls. In enzyme-linked immunosorbent assay they reacted strongly with antisera to peanut green mosaic and soybean mosaic virus antisera, and moderately with adzuki bean mosaic and peanut stripe virus antisera. All isolates also reacted positively with antisera to peanut eye spot, blackeye cowpea mosaic, pea seed-borne mosaic, potato virus Y and tobacco etch viruses, and did not react with antisera to peanut mottle, bean yellow mosaic, bean common mosaic, clover yellow vein and sugarcane mosaic viruses. SDS-PAGE analysis of purified virus preparations of the three isolates showed a single polypeptide with mol. wt. of 34,500 daltons. Based on these results, the three isolates are identified as biologically distinct strains of peanut green mosaic virus.     

Development of immunochromatographic test systems for express detection of plant viruses

Express immunochromatographic test-strip assays were developed for detection of five plant viruses varying in shape and size of virions: spherical carnation mottle virus, bean mild mosaic virus, rodshaped tobacco mosaic virus, and filamentous potato viruses X and Y. Multimembrane composites (test strips) with immobilized polyclonal antibodies against viruses and colloidal gold-conjugated antibodies were used for the analysis. The immunochromatographic test strips were shown to enable the detection of viruses both in purified preparations and in leaf extracts of infected plants with a sensitivity from 0.08 to 0.5 μg/ml for 10 min. The test strips may be used for express diagnostics of plant virus diseases in field conditions.     

Purification and Serology of the Iranian Maize Mosaic Rhabdovirus1)

Leaves of maize infected with the Iranian maize mosaic rhabdovirus (IMMRV) were homogenized in 0.1 M citrate-0.04 M Na₂SO₃ buffer, pH 5.4, containing 10 % sucrose and the extract was subjected to low speed and high speed centrifugation followed by resuspension in 0.05 M potassium phosphate buffer, pH 7.2, containing 10 % sucrose. Partially purified preparation was obtained by density-gradient centrifugation, removal of the virus zones and their concentration by high speed centrifugation. Two virus specific bands were observed in density-gradient columns. An antiserum with a titer of 128 was prepared by injecting partially purified virus into rabbits. In agar-gel-diffusion tests, the antiserum produced one or two precipitin lines against diseased maize extract but none against healthy maize extract. IMMRV was not related to barley yellow striate mosaic (BYSMV), cereal chlorotic mottle (CCMV), Cynodon chlorotic streak (CCSV), Festuca leaf streak, and maize mosaic (MMV) viruses as well as to two unidentified rhabdoviruses occurring in wheat and Bermuda grass in the vicinity of Shiraz, when these viruses were tested against IMMRV antiserum in agar-gel-diffusion and enzyme-linked immunosorbent assay. Likewise, IMMRV did not react with antisera to BYSMV, CCMV, CCSV and MMV in agar-gel-diffusion tests. IMMRV appears to be different from most reported rhabdoviruses of cereals.     

VIRUSES OF COWPEA, VIGNA UNGUICULATA L. (WALP.), IN NIGERIA

The cowpea strain of tobacco mosaic virus was isolated from a range of leguminous hosts at Ibadan, but was rare in cultivated crops. Systemic symptoms in species infected experimentally are described.
A new virus of cowpea was also found in Nigeria. The physical properties (thermal inactivation point 56° C., dilution end-point 1/50,000 and longevity in vitro 4 days at 25° C.) differ from those for cowpea viruses reported elsewhere and the name cowpea yellow mosaic virus is proposed. This virus produces local lesions in French bean ( Phaseolus vulgaris L.) and local and systemic lesions in Bengal bean ( Mucuna aterrima Holland), but does not infect other leguminous hosts. The virus was purified and an antiserum prepared against it.
Both viruses are transmitted by a beetle ( Ootheca mutabilis Sahlb.) which loses infectivity within 48 hr. of leaving plants infected with either or both viruses.     

国家种质库保存大豆和菜豆种质的种传病毒检测

以国家种质中期库提供的300份大豆、100份菜豆种质为材料,分别采用血清学和分子生物学方法,对种传病毒的种类进行了检测。结果表明:在大豆种质中检测出大豆花叶病毒(SMV)、黄瓜花叶病毒(CMV)、苜蓿花叶病毒(AMV)3种病毒,阳性检出率分别为25.33%(76份)、13.67%(41份)和4.67%(14份)。大豆种质中还存在大豆花叶病毒与黄瓜花叶病毒、大豆花叶病毒与苜蓿花叶病毒的复合侵染。在菜豆种质中检测出菜豆普通花叶病毒(BCMV)阳性材料92份,种质带毒率高达92%。这些信息将会对今后采取相关措施提高国家种质库保存的豆类种质的质量提供帮助。     

The Use of a Simple Electron Microscope Serology Procedure to Observe Relationships of Seven Potyviruses

The relationships between bean yellow mosaic (BYMV), bean common mosaic (BCMV), clover yellow vein (CYVV), lettuce mosaic (LMV), potato virus Y (PVY), turnip mosaic (TuMV) and celery mosaic (CeMV) viruses were studied in homologous and heterologous reactions, using simple and relatively rapid electron microscope serology decoration tests. The degree of relationship between these viruses was assessed by the intensity of antibody coating when the viruses were decorated by heterologous antibodies. A close relationship was observed between BYMV and CYVV, and between BYMV and LMV but not between CYVV and LMV. CeMV was quite closely related to BYMV and CYVV. Antibodies to BCMV and BYMV intensely decorated different strains of their own virus, but decoration was negligible in cross reactions.     

Serological diagnosis and host range studies of important viral diseases of a few cucurbitaceous crops in Maharashtra,India

Around 39 well characterised viruses affect cucurbits crops in developing countries and their viral diversity may be the consequence for genetic and ecological diversity of their hosts. Indeed, cucurbits are grown in variety of climatic, environmental and agricultural conditions, and this may provide more or less favourable conditions for the specific viruses or their hosts. The presence of various viral diseases caused by different viruses in Maharashtra was studied from the infected samples collected from cucurbits and melons during the survey conducted in 2009–2010 in different locations. The virus isolates collected from various cucurbitaceous crops were established and their host ranges were studied by sap transmission. The study revealed Cucumber Mosaic Virus (CMV), Zucchini yellow mosaic virus (ZYMV), Watermelon mosaic virus (WMV) and Cucumber green mottle mosaic virus infections predominately found in Nashik region, and Watermelon bud necrosis virus (WBNV), CMV, ZYMV, WMV and Watermelon silver mottle virus (WSMoV) infections in Aurangabad and Paithan regions. In Sangamner region, the crop was mostly affected by WBNV, ZYMV and WSMoV, and CMV was found only in Sillod region. The protocols for performing sap transmission tests in assay hosts were standardised for ZYMV, CMV and WBNV. Using direct antigen-coating enzyme-linked immunosorbent assay, of all the plant parts, young leaves were found to have high concentration of virus and suitable for virus detection in screening programmes. CMV and ZYMV was found to have high concentration of virus and suitable for virus detection in screening programmes.