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1.
Bernd Reiss  Rolf Sprengel  Hans Will  Heinz Schaller   《Gene》1984,30(1-3):211-217
A general method is described for the detection and quantification of low amounts of neomycin phosphotransferase in crude cell extracts. The assay is based on the electrophoretic separation of the enzyme from other interfering proteins and detection of its enzymatic activity by in situ phosphorylation of the antibiotic kanamycin. Both kanamycin and [γ32P]ATP acting as substrates are embedded in an agarose gel placed on the polyacrylamide gel containing the separated proteins. After the enzymatic reaction, the phosphorylated kanamycin is transferred to P81 phosphocellulose ion exchange paper and the radiolabeled kanamycin is visualised by autoradiography. With this method 1 ng of active enzyme can easily be detected. Both prokaryotic and eukaryotic cell extracts can be examined, and changes in the size of enzymatically active proteins can be determined.  相似文献
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本文通过对小麦蜡质蛋白的实验和研究,详细论述了蜡质蛋白的特性、提取 原理、电泳原理和检验原理。对多倍体麦类作物Wx蛋白亚基的检测,提出了应用效果良好的电泳系统,并就提取方法、凝胶配方和操作步骤进行了描述。本文还列出了部分麦类作物蜡质蛋白亚基基本模式示意图和实验例证,具有很好的参考价值。 Abstract:The waxy protein property,principles of its extraction,electrophoresis and detection were discussed in detail in this paper.According to the detection of polyploid wheat waxy subunits,an effective electrophoresis system was given with the extraction method,gel composition and operating steps listed.The basic iso-form patterns of wheat and related species as well as an application example were also showed in the paper.It is recommended that this paper is valuable both as reference and in application.  相似文献
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DNA or 2-deoxyguanosine reacts with hydroxyl free radical to form 8-hydroxy-deoxyguanosine (8-OH-dG). We found that 8-OH-dG can be effectively separated from deoxyguanosine by high pressure liquid chromatography and very sensitively detected using electrochemical detection. The sensitivity by electrochemical detection is about one-thousand fold enhanced over optical detection. Utilizing deoxyguanosine in bicarbonate buffer it was found that ferrous ion, but not ferric ion, was effective in forming 8-OH-dG. The hydroxyl free radical scavenging agents, thiourea and ethanol, were very effective in quenching Fe(11) mediated 8-OH-dG formation, but superoxide dismutase had very little effect.  相似文献
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The aim of the present work was to study colonization patterns in roots by different arbuscular mycorrhizal fungi developing from a mixed community in soil. As different fungi cannot be distinguished with certainty in planta on the basis of fungal structures, taxon-discriminating molecular probes were developed. The 5' end of the large ribosomal subunit containing the variable domains D1 and D2 was amplified by PCR from Glomus mosseae (BEG12), G. intraradices (LPA8), Gigaspora rosea (BEG9) and Scutellospora castanea (BEG1) using newly designed eukaryote-specific primers. Sequences of the amplification products showed high interspecies variability and PCR taxon-discriminating primers were designed to distinguish between each of these four fungi. A nested PCR, using universal eukaryotic primers for the first amplification and taxon-discriminating primers for the second, was performed on individual trypan blue-stained mycorrhizal root fragments of onion and leek, and root colonization by four fungi inoculated together in a microcosm experiment was estimated. More than one fungus was detected in the majority of root fragments and all four fungi frequently co-existed within the same root fragment. Root colonization by G. mosseae and G. intraradices was similar from individual and mixed inoculum, whilst the frequency of S. castanea and Gig. rosea increased in the presence of the two Glomus species, suggesting that synergistic interactions may exist between some arbuscular mycorrhizal fungi.  相似文献
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Martingale-based residuals for survival models   总被引:27,自引:0,他引:27  
7.
副溶血弧菌LAMP检测方法的建立   总被引:26,自引:0,他引:26       下载免费PDF全文
副溶血弧菌(Vibrio parahaemolyticus)是一种能引起食源性疾病的重要病原菌。首次将一种新颖的核酸扩增技术-环介导等温扩增技术(Loop-Mediated Isothermal Amplification, LAMP)应用于副溶血弧菌的快速检测。针对副溶血弧菌不耐热溶血毒素基因(tlh)设计四条特异性引物(两条内引物和两条外引物)进行LAMP扩增,对扩增反应进行优化,最佳反应时间为60 min,反应温度为60 ℃。对12种细菌共28株菌进行LAMP扩增,仅14株副溶血弧菌得到阳性扩增结果,证明引物具有很高的特异性。副溶血弧菌基因组DNA和纯培养物的检测灵敏度分别约为90 fg和24 cfu/mL。对模拟食品样品进行直接检测,检测限为89 cfu/g。结果表明,该方法检测副溶血弧菌特异性强、灵敏度高,并且操作简便、检测成本低,1 h即可完成,有望发展成为快速检测副溶血弧菌的有效手段。  相似文献
8.
For most cancers, survival rates depend on the early detection of the disease. So far, no biomarkers exist to cope with this difficult task. New proteomic technologies have brought the hope of discovering novel early cancer-specific biomarkers in complex biological samples and/or of the setting up of new clinically relevant test systems. Novel mass spectrometry-(MS) based technologies in particular, such as surface-enhanced laser desorption/ionisation time of flight (SELDI-ToF-MS), have shown promising results in the recent literature. Here, proteomic profiles of control and disease states are compared to find biomarkers for diagnosis. This paper aims to address the authors' own work and that of other groups in clinical cancer proteomics based on SELDI-ToF-MS. Shortcomings and hopes for the future are discussed.  相似文献
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转基因植物食品检测技术研究进展   总被引:24,自引:0,他引:24  
随着转基因植物的迅速发展,转基因植物(GMO)含量大量涌向市场。出于对人类健康的考虑,GMO食品标识制度在世界范围内受到人们普遍关注。不论是对GMO食品贴标签,或是对GMO与非GMO原料的分别输送,GMO原料和食品的检测技术是必不可少的。GMO食品的检测主要有两种方法:一种是以DNA为基础的PCR法,另一种是从蛋白质水平出的ELISA法。比较全面地阐述了这两种检测方法的应用状况,重点介绍了PCR检测法及影响PCR检测方法的因素,作者预测DNA芯片检测法将是未来的发展方向。  相似文献
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