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Detection of Colletotrichum coccodes from soil and potato tubers by conventional and quantitative real-time PCR 总被引:4,自引:1,他引:4
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 . 相似文献
3.
D. Sivakumar R. S. Wilson Wijeratnam M. Abeyesekere R. L. C. Wijesundera 《Phytoparasitica》2002,30(1):43-51
Botryodiplodia theobromae, Colletotrichum gloeosporioides andGliocephalotrichum microchlamydosporum are the causal fungi of the rambutan postharvest diseases stem-end rot, anthracnose and brown spot, respectively. Two different
treatments of rambutan fruits(Nephelium lappaceum) against the three pathogens were compared: potassium metabisulphite (250 ppm) or cinnamaldehyde (30 ppm), each combined withTrichoderma harzianum (TrH 40). The application of TrH 40 and potassium metabisulphite effectively controlled the incidence and severity of the
three postharvest diseases and maintained the overall quality and color of the fruit under low temperature storage at 13.5°C
and 95% r.h. for 18 days. The greatest effect of this treatment was shown onG. microchlamydosporum. Cinnamaldehyde affected the growth and germination of TrH 40, whereas potassium metabisulphite did not.
http://www.phytoparasitica.org posting Nov. 4, 2001. 相似文献
4.
辣椒疫病发病主导因素及综合防治技术研究 总被引:2,自引:0,他引:2
通过3a在多年辣椒重茬地的田间试验及调查研究,基本明确了辣椒疫病发生与灌水方法、栽培方式、降水、中心病株的出现等主导因素的关系。提出了栽培防病、化学治疗的综防技术,达到控病增产的目的。 相似文献
5.
Seiju Ishikawa 《Journal of General Plant Pathology》2004,70(5):249-255
Simple diagnosis by ethanol immersion (SDEI) to detect Glomerella cingulata was used to detect three other fungi that also cause latent infection of strawberry plants. Signs on strawberry leaves with asymptomatic latent infection by Colletotrichum acutatum became visible using SDEI. Salmon-pink conidial masses were produced in the acervuli on the treated leaves 5 days after incubation at 28°C. In the case of Dendrophoma obscurans, pycnidia with amber conidial masses formed 5 days after incubation at 28°C. The pycnidia were observed mainly on the ribs, and conidial masses exuded from the ostiole. These macroscopic conidial masses were similar to those of G. cingulata and C. acutatum. When water was dripped onto a lesion caused by D. obscurans, the pycnidia exuded white filamentous conidial masses, making the distinction of D. obscurans from G. cingulata or C. acutatum. On petioles with latent infection by Fusarium oxysporum f. sp. fragariae, white aerial hyphae grew out from the vascular tissues on the cut surface 3 days after incubation at 28°C and were easily observed by eye or with a loupe. Thus, SDEI was also useful for diagnosing latent infection of strawberry plants by C. acutatum, D. obscurans, and F. oxysporum f. sp. fragariae. 相似文献
6.
辣椒疫霉是一种侵染性植物病原,能引起多种茄科及葫芦科植物的病害。利用同源克隆法从辣椒疫霉基因组内克隆了漆酶基因Pclac3,该基因全长1 881个核苷酸,编码626个氨基酸,与已知的真菌漆酶基因具有较高的同源性,并且具备漆酶基因的保守区域。利用PCR将该基因克隆于pPIC9K载体,并转化于毕赤酵母GS115,经过1%甲醇诱导表达获得其异源表达产物。将表达产物进行SDS-PAGE检测,获得分子量大约为90 kDa的特异蛋白。利用ABTS法对Pclac3的表达产物粗酶液进行酶活性分析发现,其活性在第11天时最大,达到45 U/mL。这为研究辣椒疫霉漆酶基因家族及探索卵菌漆酶生物活性及其潜在的应用提供了理论依据。 相似文献
7.
为了获得小叶九里香枝干中抑制炭疽病菌的活性成分,在活性追踪指导下,利用硅胶柱色谱、Sephadex LH-20凝胶柱色谱及半制备液相色谱等方法对小叶九里香枝干进行了研究。经追踪分离鉴定到1个活性化合物XY-01,利用质谱、核磁共振谱鉴定化合物为九里香甲碱(Koenidine),该化合物可有效抑制香蕉炭疽病菌和芒果炭疽病菌的生长,EC50值分别为34.28和45.88μg/mL,表明九里香甲碱抑菌活性较好、结构简单,具有开发为新型植物源杀菌剂的潜在价值,九里香甲碱系首次从小叶九里香中分离得到。 相似文献
8.
为了分离和克隆辣椒中疫霉菌诱导基因,以接种辣椒疫霉菌的叶片为材料,利用SMART技术构建辣椒疫霉菌与辣椒互作的双杂交c DNA文库。结果表明:该文库容量为3.6×106cfu,重组率88%左右,插入片段集中在300~2000bp之间,平均长度约为800bp,表明获得的文库质量较高,该文库将为分子育种提供重要的基因资源。 相似文献
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