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961.
Aims: To optimize and evaluate fluorescence microscopy assays for specific assessment of mycobacteria and co‐contaminants, including culturable and non‐culturable sub‐populations, in metalworking fluids (MWF). Methods and Results: Auramine‐O‐rhodamine (AR) staining and LIVE/DEAD BacLight? Bacterial Viability staining (L/D staining) were adapted and evaluated for detection/quantification and differentiation (viable vs non‐viable) of the MWF‐associated mycobacteria and the background bacterial flora, respectively. The AR staining method was found to be specific to MWF mycobacteria with a minimum detection limit of 10 cells ml?1 and was comparable to the QPCR in quantification efficiency in MWF matrix. The L/D staining‐based microscopy allowed differential quantification of viable vs non‐viable cells. In general, a 3‐log difference was observed between the L/D microscopy count and culture count accounting for the presence of non‐culturable fraction in the bacterial population in in‐use MWF. Conclusions: The optimized AR staining‐ and the L/D staining‐based microscopy methods have the potential for rapid, specific and differential assessment (viable vs non‐viable) of MWF‐associated mycobacteria and co‐contaminants in field MWF. Significance and Impact of the study: Early detection of MWF mycobacteria by rapid, low‐cost, less‐skill intensive and culture‐independent fluorescence‐based microscopy methods will facilitate timely intervention to protect the machine workers from occupational hazards. 相似文献
962.
AIMS: To investigate the difference between Lancefield group C Streptococcus dysgalactiae (GCSD) strains isolated from diseased fish and animals by sequencing and phylogenetic analysis of the sodA gene. METHODS AND RESULTS: The sodA gene of Strep. dysgalactiae strains isolated from fish and animals were amplified and its nucleotide sequences were determined. Although 100% sequence identity was observed among fish GCSD strains, the determined sequences from animal isolates showed variations against fish isolate sequences. Thus, all fish GCSD strains were clearly separated from the GCSD strains of other origin by using phylogenetic tree analysis. In addition, the original primer set was designed based on the determined sequences for specifically amplify the sodA gene of fish GCSD strains. The primer set yield amplification products from only fish GCSD strains. CONCLUSIONS: By sequencing analysis of the sodA gene, the genetic divergence between Strep. dysgalactiae strains isolated from fish and mammals was demonstrated. Moreover, an original oligonucletide primer set, which could simply detect the genotype of fish GCSD strains was designed. SIGNIFICANCE AND IMPACT OF THE STUDY: This study shows that Strep. dysgalactiae isolated from diseased fish could be distinguished from conventional GCSD strains by the difference in the sequence of the sodA gene. 相似文献
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964.
目的建立快速、灵敏、特异的分子生物学检测卫氏并殖吸虫的方法。方法根据卫氏并殖吸虫的特异性基因序列,设计适合于PCR检测的特异性引物及实时荧光PCR特异性引物和探针,并进行灵敏性和特异性试验。结果设计的引物和探针特异性强,所建立的检测方法灵敏度高。应用实时荧光PCR方法的检测灵敏度可达到0.1 pg/μL,比PCR方法的灵敏度高三个数量级。结论本研究所建立的PCR和实时荧光PCR技术检测卫氏并殖吸虫方法的特异性强,灵敏度高,为卫氏并殖吸虫感染的诊治提供了快速的检测技术手段。 相似文献
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968.
目的探讨Kruppel样因子4(Kruppel-like factor 4,KLF4)在内毒素血症小鼠中的表达模式及意义。方法运用实时荧光PCR技术和Western blot技术,分别从mRNA水平和蛋白水平探讨内毒素血症小鼠肝脏和肺脏中KLF4的表达;运用生物信息学技术,对启动子区含有KLF4的结合位点的炎症介质基因进行了预测;运用RT-PCR技术,从mRNA水平探讨内毒素血症小鼠肝脏和肺脏中IL1β的表达模式。结果内毒素血症小鼠肝脏和肺脏中KLF4 mRNA的表达下凋,KLF4蛋白的表达先下凋后升高;IL-18、IL-15、IL-12、IL-18、IL-10等炎症介质基因的启动子区均含有KLF4的结合元件,这些炎症基因的表达可能直接受到KLF4的调控;内毒素血症小鼠肝脏和肺脏中IL-IB的表达模式与KLF4的表达模式呈相反趋势。结论内毒素血症小鼠肝脏和肺脏中KLF4表达下调,KLF4在炎症介质基因表达调控中可能具有重要作用。 相似文献
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970.
Sa-nguanmoo P Thongmee C Ratanakorn P Pattanarangsan R Boonyarittichaikij R Chodapisitkul S Theamboonlers A Tangkijvanich P Poovorawan Y 《Journal of medical primatology》2008,37(6):277-289
Background Hepatitis B virus (HBV) is a public health problem worldwide and apart from infecting humans, HBV has been found in non‐human primates. Methods We subjected 93 non‐human primates comprising 12 species to ELISA screening for the serological markers HBsAg, antiHBs and antiHBc. Subsequently, we detected HBV DNA, sequenced the whole HBV genome and performed phylogenetic analysis. Results HBV infection was detected in gibbon (4/15) and orangutan (7/53). HBV DNA isolates from two gibbons and seven orangutans were chosen for complete genome amplification. We aligned the Pre‐S/S, Pre‐C/C and entire genomes with HBV sequences and performed phylogenetic analysis. The gibbon and orangutan viruses clustered within their respective groups. Conclusions Both geographic location and host species influence which HBV variants are found in gibbons and orangutans. Hence, HBV transmission between humans and non‐human primates might be a distinct possibility and additional studies will be required to further investigate this potential risk. 相似文献