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51.
Barnaby Kelly Douglas Thamm Rhonda J. Rosengren 《Veterinary and comparative oncology》2023,21(4):595-604
Canine osteosarcoma is an aggressive cancer, comprising 85% of canine bone neoplasms. Current treatment practices of surgery and chemotherapy increase 1-year survival by only 45%. The curcumin analogue RL71, has demonstrated potent in vitro and in vivo efficacy in several models of human breast cancer through increased apoptosis and cell cycle arrest. Thus, the present study aimed to investigate efficacy of curcumin analogues in two canine osteosarcoma cell lines. Osteosarcoma cell viability was assessed using the sulforhodamine B assay and mechanisms of action were determined by analysing the levels of cell cycle and apoptotic regulatory proteins via Western blotting. Further evidence was obtained using flow cytometry to detect cell cycle distribution and the number of apoptotic cells. RL71 was the most potent curcumin analogue with EC50 values of 0.64 ± 0.04 and 0.38 ± 0.009 μM (n = 3) in D-17 (commercial) and Gracie canine osteosarcoma cells, respectively. RL71 significantly increased the ratio of cleaved-caspase 3 to pro-caspase 3 and the level of apoptotic cells at the 2× and 5× EC50 concentration (p < 0.001, n = 3). Furthermore, at the same concentration, RL71 significantly increased the number of cells in the G2/M phase. In conclusion, RL71 has potent cytotoxic activity in canine osteosarcoma cells triggering G2/M arrest and apoptosis at concentrations achievable in vivo. Future research should further investigate molecular mechanisms for these changes in other canine osteosarcoma cell lines prior to in vivo investigation. 相似文献
52.
单细胞转录组测序(single-cell RNA sequencing,scRNA-seq)技术在单细胞水平对转录组进行测序,可从表达量、细胞量及细胞组成等多种角度描述样本,主要包括单细胞分离、mRNA反转录、文库构建、转录组测序和数据分析等步骤。单细胞分离是scRNA-seq技术操作的第一步,常用的细胞分离方法有连续稀释法、显微操作法、荧光激活流式细胞术、激光捕获显微切割术和微流控技术。基于不同细胞捕获、cDNA扩增和文库构建开发了许多scRNA-seq方法,如CEL-seq2、Drop-seq、MARS-seq、SCRB-seq、Smart-seq、Smart-seq2等。与传统测序方法相比,scRNA-seq可识别单个细胞的基因表达信息、记录细胞的空间位置、跟踪不同细胞谱系在分化过程中的轨迹,这为研究难以大量提取的肌内脂肪细胞分子特异性和发生分化过程带来极大便利。作者简要介绍了scRNA-seq技术的关键步骤,比较分析了单细胞分离和转录组测序方法的优缺点,综述了肌内脂肪细胞的起源以及scRNA-seq在肌内脂肪细胞的亚群和标记基因鉴定以及细胞分化轨迹追踪中的应用,以期为肌内脂肪细胞的深入研究提供参考依据。 相似文献
53.
54.
观察miR-21与Nocodazole联合作用对小鼠成肌细胞C2C12周期的影响。分别以0、200、300、400、500、600 nmol/L Nocodazole处理C2C12细胞24 h,流式细胞仪检测细胞周期,间接免疫染色激光共聚焦显微镜观察细胞有丝分裂器α-微管蛋白(α-tubulin)排列。转染miR-21 mimics/NC和inhibitors/NC,24 h后,添加400 nmol/L的Nocodazole处理24 h,流式细胞仪检测细胞周期。结果表明:Nocodazole使C2C12细胞同步化的最佳浓度是400 nmol/L;过表达miR-21 mimics之后,与对照组相比,G0/G1,S期细胞百分比极显著增加,G2/M期细胞百分比极显著减少(P0.01);添加抑制剂后,与对照组相比,添加抑制剂(inhibitors)的C2C12细胞中处于G0/G1期的细胞比例显著高于对照组,而处于G2/M期的细胞比例显著低于对照组细胞(P0.05);正反试验结果证明,miR-21促进C2C12细胞周期进入S期。为进一步研究miR-21对C2C12细胞的作用机制奠定基础。 相似文献
55.
《Veterinary microbiology》2015,175(2-4):211-217
Squamous cell carcinomas are common feline skin cancers that have been associated with infection with Felis catus papillomavirus type 2 (FcaPV-2). Currently, little is known about the epidemiology of FcaPV-2 infection. The aim of this study was to develop a real-time PCR assay to quantify FcaPV-2 DNA in plucked hairs and skin swabs from 11 healthy breeding queens and their kittens. Samples were taken prior to kittening and then 2, 7 and 28 days after kittening to determine the age at which the kittens were first exposed to the virus. FcaPV-2 DNA was amplified from all of the queens and from 91% of the kittens at 2 days of age. There was a wide range in the quantity of FcaPV-2 DNA detected, from 1 to 92,520 copies per swab, and from 0.01 to 234 copies per copy of reference gene DNA in the hair plucks. The quantity of FcaPV-2 DNA detected in samples collected from the kittens was strongly correlated to that of their respective queens and the mean viral DNA load was similar for cats within a household but varied significantly between households. This is the first time that quantitative PCR has been used to detect FcaPV-2 DNA and the results suggest that the virus is ubiquitous but there is a wide variation of viral DNA loads. Kittens appear to be exposed to FcaPV-2 early in life, presumably from direct contact with their queen. These results are important when determining if FcaPV-2 infection of cats is preventable. 相似文献
56.
Anna Kakehashi Akihiro Hagiwara Norio Imai Min Wei Shoji Fukushima Hideki Wanibuchi 《Journal of toxicologic pathology》2015,28(1):27-32
In the present study, in continuation of our previous experiment in order to investigate the mode of action (MOA) of ethyl tertiary-butyl ether (ETBE) hepatotumorigenicity in rats, we aimed to examine alterations in cell proliferation, that are induced by short-term administration of ETBE. F344 rats were administered ETBE at doses of 0, and 1,000 mg/kg body weight twice a day by gavage for 3, 10, 17 and 28 days. It was found that the previously observed significant increase of P450 total content and hydroxyl radical levels after 7 days of ETBE administration, and 8-OHdG formation at day 14, accompanied by accumulation of CYP2B1/2B2, CYP3A1/3A2, CYP2C6, CYP2E1 and CYP1A1 and downregulation of DNA oxoguanine glycosylase 1, was preceded by induction of cell proliferation at day 3. Furthermore, we observed an increase in regenerative cell proliferation as a result of ETBE treatment at day 28, followed by induction of cell cycle arrest and apoptosis by day 14. These results indicated that short-term administration of ETBE led to a significant early increase in cell proliferation activity associated with induction of oxidative stress, and to a regenerative cell proliferation as an adaptive response, which could contribute to the hepatotumorigenicity of ETBE in rats. 相似文献
57.
Rei NAKANO Kazuya EDAMURA Tomohiro NAKAYAMA Kenji TESHIMA Kazushi ASANO Takanori NARITA Ken OKABAYASHI Hiroshi SUGIYA 《The Journal of veterinary medical science / the Japanese Society of Veterinary Science》2015,77(1):27-35
We investigated the in
vitro differentiation of canine bone marrow stromal cells (BMSCs) into voltage-
and glutamate-responsive neuron-like cells. BMSCs were obtained from the bone marrow of
healthy beagle dogs. Canine BMSCs were incubated with the basal medium for neurons
containing recombinant human basic fibroblast growth factor (bFGF; 100
ng/ml). The viability of the bFGF-treated cells was
assessed by a trypan blue exclusion assay, and the morphology was monitored. Real-time
RT-PCR was performed to evaluate mRNA expression of neuronal, neural stem cell and glial
markers. Western blotting and immunocytochemical analysis for the neuronal markers were
performed to evaluate the protein expression and localization. The Ca2+
mobilization of the cells was evaluated using the Ca2+ indicator Fluo3 to
monitor Ca2+ influx. To investigate the mechanism of bFGF-induced neuronal
differentiation, the fibroblast growth factor receptor inhibitor, the phosphoinositide
3-kinase inhibitor or the Akt inhibitor was tested. The bFGF treatment resulted in the
maintenance of the viability of canine BMSCs for 10 days, in the expression of neuronal
marker mRNAs and proteins and in the manifestation of neuron-like morphology. Furthermore,
in the bFGF-treated BMSCs, a high concentration of KCl and L-glutamate induced an increase
in intracellular Ca2+ levels. Each inhibitor significantly attenuated the
bFGF-induced increase in neuronal marker mRNA expression. These results suggest that bFGF
contributes to the differentiation of canine BMSCs into voltage- and glutamate-responsive
neuron-like cells and may lead to the development of new cell-based treatments for
neuronal diseases. 相似文献
58.
Fong-Yuan LIN Yeu-Yang TSENG Kun-Wei CHAN Shu-Ting KUO Cheng-Hsiung YANG Chi-Young WANG Masaki TAKASU Wei-Li HSU Min-Liang WONG 《The Journal of veterinary medical science / the Japanese Society of Veterinary Science》2015,77(9):1055-1062
Orf virus (ORFV), a member of parapoxvirus, is an enveloped virus with genome
of double-stranded DNA. ORFV causes contagious pustular dermatitis or contagious ecthyma
in sheep and goats worldwide. In general, detection of viral DNA and observing ORFV virion
in tissues of afflicted animals are two methods commonly used for diagnosis of orf
infection; however, isolation of the ORFV in cell culture using virus-containing tissue as
inoculum is known to be difficult. In this work, the ORFV (Hoping strain) isolated in
central Taiwan was successfully grown in cell culture. We further examined the biochemical
characteristic of our isolate, including viral genotyping, viral mRNA and protein
expression. By electron microscopy, one unique form of viral particle from ORFV infected
cellular lysate was demonstrated in the negative-stained field. Moreover, immunomodulating
and anti-influenza virus properties of this ORFV were investigated. ORFV stimulated human
monocytes (THP-1) secreting proinflammatory cytokines IL-8 and TNF-α. And, pre-treatment
of ORFV-infected cell medium prevents A549 cells from subsequent type A influenza virus
(IAV) infection. Similarly, mice infected with ORFV via both intramuscular and
subcutaneous routes at two days prior to IAV infection significantly decreased the
replication of IAV. In summary, the results of a current study indicated our Hoping strain
harbors the immune modulator property; with such a bio-adjuvanticity, we further proved
that pre-exposure of ORFV protects animals from subsequent IAV infection. 相似文献
59.
为探究高效氯氰菊酯 (beta-cypermethrin, β-CP) 对雌性小鼠卵巢生殖功能的影响及维生素E (vitamin E, VE) 的干预作用,将雌性昆白小鼠随机分为6 组:空白对照组 (花生油处理)、β-CP不同剂量 (10、20、40 mg/kg) 处理组、VE保护组 (20 mg/kg β-CP+20 mg/kg VE) 和VE组 (20 mg/kg VE),连续灌胃10 d。灌胃结束后取小鼠卵巢组织,观察组织结构的病理变化,采用免疫组化法、蛋白免疫印迹试验及RT-PCR方法检测卵巢中StAR蛋白含量及casp-3、casp-8、INHα和INHβB 基因mRNA表达的变化。结果显示:与对照相比,10、20和40 mg/kg的β-CP处理均使小鼠卵巢组织结构发生了损伤,使组织中StAR蛋白的浓度分别降低了18.8%、36.3%和40.3%,casp-3基因的表达分别升高了16.0%、26.7%和52.9%,INHα基因的表达分别升高了34.5%、83.6%和228.7%,INHβB基因的表达分别升高了7.5%、39.2%和52.7%;20和40 mg/kg的β-CP处理使得casp-8基因的表达分别升高了27.1%和36.7%。上述处理组与对照组的差异均达显著水平 (P<0.05)。与 20 mg/kg β-CP处理组相比,VE 保护组的StAR蛋白含量也显著增多 (P<0.05)。研究表明,β-CP对小鼠卵巢具有毒性作用,这与β-CP抑制StAR蛋白的合成,上调casp-3、casp-8、INHα及INHβB基因的表达有关;添加VE对小鼠卵巢有一定的保护作用,这与VE可减弱β-CP对StAR蛋白合成的抑制作用有关。 相似文献
60.
Inoculation of wheat(Triticum aestivum L.) leaves with wheat powdery mildew fungus(Blumeria graminis f. sp. tritici) induces the cell death in adventitious roots. Reactive oxygen species(ROS) play a key role in respond to biotic stress in plants. To study the involvement of ROS and the degree of cell death in the wheat roots following inoculation, ROS levels and microstructure of root cells were analyzed in two wheat cultivars that are susceptible(Huamai 8) and resistant(Shenmai 8) to powdery mildew fungus. At 18 d after powdery mildew fungus inoculation, only Huamai 8 displayed the leaf lesions, while root cell death occurred in both varieties. Huamai 8 had a high level of ROS accumulation, which is associated with increased root cell degradation, while in Shenmai 8, there was little ROS accumulation correlating with slight root cell degradation. The molecular study about the expression levels of ROS scavenging genes(MnSOD and CAT) in wheat roots showed that these genes expression decreased after the leaves of wheat was inoculated. The difference between Huamai 8 and Shenmai 8 on subcellular localization of H2 O2 and O2–· was corresponded with the different down-regulation of the genes encoding for superoxide dismutase and catalase in two wheat cultivars. These results suggested that ROS were involved in the process by which powdery mildew fungus induced cell death in wheat roots. 相似文献