Engineering of a Single Chain Variable Fragment Antibody Specific for the Citrus tristeza virus and its Expression in Escherichia coli and Nicotiana tabacum

Citrus tristeza virus (CTV) is one of the most destructive citrus virus diseases in the world. The construction of an engineered antibody, EMBL accession number AJ278109, able to specifically recognize its antigen, i.e. the coat protein of CTV, directly on infected plant material without any purification or manipulation of the entire woody plant. The potential uses of this engineered antibody are discussed.     

The Complete Nucleotide Sequence of a Japanese Isolate of White clover mosaic virus Strain RC

The complete nucleotide sequence was determined for genomic RNA of White clover mosaic virus (WClMV-RC) isolated from red clover (Trifolium pratense) in Japan, It is 5843 nucleotides in length, excluding the poly(A) tail at the 3' terminus. Similar to other potexviruses, it contains five open reading frames (ORFs 1 through 5), which putatively encode an RNA-dependent RNA polymerase (RdRp) (147 kDa), a triple gene block (TGB) (26 kDa/13 kDa/7 kDa), and a coat protein (CP) (22 kDa), respectively. The deduced amino acid sequence of the WClMV-RC CP was identical to that of WClMV-O, one of two New Zealand isolates, but only 85% identical to that of WClMV-M, the other New Zealand isolate, because of heterogeneity in the C-termini of CP amino acid sequences. The implication of this CP heterogeneity is discussed. Received 30 August 2001/ Accepted in revised form 11 January 2002     

Detection of Colletotrichum coccodes from soil and potato tubers by conventional and quantitative real-time PCR

Colletotrichum coccodes is the causal agent of the potato blemish disease black dot. Two PCR primer sets were designed to sequences of the ribosomal internal transcribed spacer (ITS1 and ITS2) regions for use in a nested PCR. The genus-specific outer primers (Cc1F1/Cc2R1) were designed to regions common to Colletotrichum spp., and the species-specific nested primers (Cc1NF1/Cc2NR1) were designed to sequences unique to C . coccodes . The primer sets amplified single products of 447 bp (Cc1F1/Cc2R1) and 349 bp (Cc1NF1/Cc2NR1) with DNA extracted from 33 European and North American isolates of C. coccodes. The specificity of primers Cc1NF1/Cc2NR1 was confirmed by the absence of amplified product with DNA of other species representing the six phylogenetic groups of the genus Colletotrichum and 46 other eukaryotic and prokaryotic plant pathogenic species. A rapid procedure for the direct extraction of DNA from soil and potato tubers was used to verify the PCR assay for detecting C. coccodes in environmental samples. The limit of sensitivity of PCR for the specific detection of C. coccodes when inoculum was added to soils was 3·0 spores per g, or the equivalent of 0·06 microsclerotia per g soil, the lowest level of inoculum tested. Colletotrichum coccodes was also detected by PCR in naturally infested soil and from both potato peel and peel extract from infected and apparently healthy tubers. Specific primers and a TaqMan fluorogenic probe were designed to perform quantitative real-time (TaqMan) PCR to obtain the same levels of sensitivity for detection of C. coccodes in soil and tubers during a first-round PCR as with conventional nested PCR and gel electrophoresis. This rapid and quantitative PCR diagnostic assay allows an accurate estimation of tuber and soil contamination by C. coccodes .     

Bt对小地考虑呼吸作用的影响

利用红外CO2分析仪测定小地老虎幼虫呼吸节律的变化结果显示：小地老虎幼虫呼吸节律属于不规则波动型，取食含Bt烟叶的幼虫呼出的CO2量显著低于对照组，且呼吸节律的波动性也明显增大，说明小地考虑幼虫取食Bt后，呼吸代谢受到一定干扰，出现呼吸水平降低及代谢紊乱现象。     

晋南冬麦区大麦黄矮病毒流行株系监测及防治策略探讨

连续5年(1996～2000年)采集晋南冬麦区小麦黄矮病标样,采用生物学和血清学(酶联免疫吸附法)相结合的诊断方法对该地区的大麦黄矮病毒流行株系进行了鉴别。结果表明,该小麦黄矮病流行区近五年以GAV株系为主流株系,兼有少量GPV、PAV和混合株系存在。同时对小麦抗黄矮病新品种“临抗1号”进行了GPV和GAV两种株系的抗性测定,明确了该品种兼抗GPV和GAV两种株系。根据小麦黄矮病发生现状,提出了一套以选育推广抗耐病品种为主,以药剂防治为辅的综合防治措施。以期为当地小麦生产服务。     

梨自交不亲和强度不同品种花柱S糖蛋白含量的差异

 提取梨( Pyrus) 自交不亲和强度不同品种花柱可溶性蛋白质, 应用等电聚焦电泳法在凝胶板上分离出S 糖蛋白, 用图像解析法进行定量。结果表明, 花柱总S 糖蛋白含量因品种而异, 自交不亲和强、弱及自交亲和的品种每1 􀀁g 花柱可溶性蛋白中含总S 糖蛋白分别为􀀂1. 3 ng 、0. 7 ~ 0. 9 ng、 0. 7 ng 。不同的S 基因所表达的S 糖蛋白量差异显著, 即使是同一S 基因所对应的S 糖蛋白量也因品种而异。     

苹果梨中多酚氧化酶酶学特性的研究

 以苹果梨中的多酚氧化酶为研究对象, 对其酶的活性变化及其特性的研究表明, 苹果梨中的多酚氧化酶的最适pH 值是4. 6 , 最适温度是40 ℃; 亚硫酸氢钠、维生素C、柠檬酸对苹果梨中多酚氧化酶的抑制作用依次减弱, 硫酸钠几乎无抑制作用。不同底物和同底物不同浓度影响多酚氧化酶特性。     

核桃早实性状的RAPD分析

 用BSA (Bulked Segregant Analysis) 法获得了与核桃早实性状相关的RAPD 标记OPB208₉₀₀。混合分离群体的混合样来自早实品种辽1 的自然授粉实生后代。用220 个随机引物扩增早实和晚实混合样, 只有引物OPB208 ( 5. GTCCACACGG 3. ) 在早实混合样及其个体上扩增出了约900 bp 的特异带OPB208₉₀₀, 而在晚实混合样及其个体上无此特异带。用OPB208 为引物, 以19 个品种和3 个株系的DNA 为模板进行扩增,特异带OPB208₉₀₀在17 个早实品种中的12 个个体上出现, 而5 个晚实品种( 系) 上均无此带。     

红肉脐橙(Citrus sinensis L.)果肉中特征色素提取方法探索

红肉脐橙是目前为止惟一果肉呈粉红色或红色的脐橙,其呈色色素尚未见报道。为深入研究其果肉中特征色素的构成及在果实生长发育和贮存过程中色素含量的变化,本研究在对该色素初步定性的基础上对提取方法进行了探索。在分析红肉脐橙果肉中色素萃取液的紫外可见吸收光谱时发现,该色素有类胡萝卜素的特征吸收峰,将萃取液进行薄层层析则进一步证实其内含有番茄红素。在色素提取的前处理方式上,本研究认为冷冻干燥样品的浸提效果较真空干燥的样品稳定(变异系数小),且浸提率相差无几,所以选择冷冻干燥作为合适的样品前处理方式。色素浸提条件的正交实验结果表明,最佳浸提条件为50℃、1.5h内浸提4次,浸提的料液比为1:60。此外,浸提温度、浸提时间、浸提料液比和浸提次数对浸提率都有显著影响,影响最大的因素是浸提温度,最佳温度为50℃,随温度降低浸提率显著降低。其次是侵提时间的长短,最佳浸提时间为1.5h,延长或缩短0.5h都会使浸提率下降。浸提次数对浸提率的影响是最小的。     

Spatial dependence and the relationship of soil organic carbon and soil moisture in the Luquillo Experimental Forest,Puerto Rico

We used geo-spatial statistical techniques to examine the spatial variation and relationship of soil organic carbon (SOC) and soil moisture (SM) in the Luquillo Experimental Forest (LEF), Puerto Rico, in order to test the hypothesis that mountainous terrain introduces spatial autocorrelation and crosscorrelation in ecosystem and soil properties. Soil samples (n = 100) were collected from the LEF in the summer of 1998 and analyzed for SOC, SM, and bulk density (BD). A global positioning system was used to georeference the location of each sampling site. At each site, elevation, slope and aspect were recorded. We calculated the isotropic and anisotropic semivariograms of soil and topographic properties, as well as the cross-variograms between SOC and SM, and between SOC and elevation. Then we used four models (random, linear, spherical and wave/hole) to test the semi-variances of SOC, SM, BD, elevation, slope and aspect for spatial dependence. Our results indicate that all the studied properties except slope angle exhibit spatial dependence within the scale of sampling (200 – 1000 m sampling interval). The spatially structured variance (the variance due to the location of sampling sites) accounted for a large proportion of the sample variance for elevation (99%), BD (90%), SOC (68%), aspect (56%) and SM (44%). The ranges of spatial dependence (the distances within which parameters are spatially dependent) for aspect, SOC, elevation, SM, and BD were 9810 m, 3070 m, 1120 m, 930 m and 430 m, respectively. Cross correlograms indicate that SOC varies closely with elevation and SM depending on the distances between samples. The correlation can shift from positive to negative as the separation distance increases. Larger ranges of spatial dependence of SOC, aspect and elevation indicate that the distribution of SOC in the LEF is determined by a combination of biotic (e.g., litterfall) and abiotic factors (e.g., microclimate and topographic features) related to elevation and aspect. This demonstrates the importance of both elevation and topographic gradients in controlling climate, vegetation distribution and soil properties as well as the associated biogeochemical processes in the LEF.This revised version was published online in May 2005 with corrections to the Cover Date.