全文获取类型
收费全文 | 6147篇 |
免费 | 482篇 |
国内免费 | 502篇 |
专业分类
7131篇 |
出版年
2024年 | 23篇 |
2023年 | 111篇 |
2022年 | 221篇 |
2021年 | 369篇 |
2020年 | 273篇 |
2019年 | 268篇 |
2018年 | 288篇 |
2017年 | 224篇 |
2016年 | 313篇 |
2015年 | 424篇 |
2014年 | 512篇 |
2013年 | 518篇 |
2012年 | 533篇 |
2011年 | 491篇 |
2010年 | 309篇 |
2009年 | 293篇 |
2008年 | 325篇 |
2007年 | 265篇 |
2006年 | 229篇 |
2005年 | 221篇 |
2004年 | 159篇 |
2003年 | 118篇 |
2002年 | 107篇 |
2001年 | 68篇 |
2000年 | 62篇 |
1999年 | 74篇 |
1998年 | 49篇 |
1997年 | 31篇 |
1996年 | 33篇 |
1995年 | 41篇 |
1994年 | 24篇 |
1993年 | 23篇 |
1992年 | 24篇 |
1991年 | 14篇 |
1990年 | 17篇 |
1989年 | 13篇 |
1988年 | 14篇 |
1987年 | 13篇 |
1986年 | 11篇 |
1985年 | 9篇 |
1984年 | 3篇 |
1983年 | 4篇 |
1982年 | 1篇 |
1981年 | 3篇 |
1979年 | 5篇 |
1978年 | 1篇 |
排序方式: 共有7131条查询结果,搜索用时 15 毫秒
41.
Ronny Martinez Felix Jakob Ran Tu Petra Siegert Karl‐Heinz Maurer Ulrich Schwaneberg 《Biotechnology and bioengineering》2013,110(3):711-720
Bacillus gibsonii Alkaline Protease (BgAP) is a recently reported subtilisin protease exhibiting activity and stability properties suitable for applications in laundry and dish washing detergents. However, BgAP suffers from a significant decrease of activity at low temperatures. In order to increase BgAP activity at 15°C, a directed evolution campaign based on the SeSaM random mutagenesis method was performed. An optimized microtiter plate expression system in B. subtilis was established and classical proteolytic detection methods were adapted for high throughput screening. In parallel, the libraries were screened for increased residual proteolytic activity after incubation at 58°C. Three iterative rounds of directed BgAP evolution yielded a set of BgAP variants with increased specific activity (Kcat) at 15°C and increased thermal resistance. Recombination of both sets of amino acid substitutions resulted finally in variant MF1 with a 1.5‐fold increased specific activity (15°C) and over 100 times prolonged half‐life at 60°C (224 min compared to 2 min of the WT BgAP). None of the introduced amino acid substitutions were close to the active site of BgAP. Activity‐altering amino acid substitutions were from non‐charged to non‐charged or from sterically demanding to less demanding. Thermal stability improvements were achieved by substitutions to negatively charged amino acids in loop areas of the BgAP surface which probably fostered ionic and hydrogen bonds interactions. Biotechnol. Bioeng. 2013; 110: 711–720. © 2012 Wiley Periodicals, Inc. 相似文献
42.
已被发现和命名的白介素有19种,其相应的受体大多也已被克隆及鉴定。白介素及其受体的结构特征决定了其信号转导和功能效应的共性和特性。许多白介素及可溶性受体在临床疾病的治疗中可能起重要作用,同时这也带动了对白介素及其受体的基因工程研究。随着基因组计划的日趋完成和一些新的分子生物学技术方法的建立,对白介素及其受体的研究在许多方面都面临着新的机遇和挑战。 相似文献
43.
Min Lin Ying Zhao ShuQi Wang Ming Liu ZhenFeng Duan YongMei Chen Fei Li Feng Xu TianJian Lu 《Biotechnology advances》2012
Lanthanide (Ln)-doped upconversion nanoparticles (UCNPs) with appropriate surface modification can be used for a wide range of biomedical applications such as bio-detection, cancer therapy, bio-labeling, fluorescence imaging, magnetic resonance imaging and drug delivery. The upconversion phenomenon exhibited by Ln-doped UCNPs renders them tremendous advantages in biological applications over other types of fluorescent materials (e.g., organic dyes, fluorescent proteins, gold nanoparticles, quantum dots, and luminescent transition metal complexes) for: (i) enhanced tissue penetration depths achieved by near-infrared (NIR) excitation; (ii) improved stability against photobleaching, photoblinking and photochemical degradation; (iii) non-photodamaging to DNA/RNA due to lower excitation light energy; (iv) lower cytotoxicity; and (v) higher detection sensitivity. Ln-doped UCNPs are therefore attracting increasing attentions in recent years. In this review, we present recent advances in the synthesis of Ln-doped UCNPs and their surface modification, as well as their emerging applications in biomedicine. The future prospects of Ln-doped UCNPs for biomedical applications are also discussed. 相似文献
44.
为探讨柘叶Cudrania tricuspidata饲养家蚕Bombyx mori易感染核多角体病毒(BmNPV)的机制, 本实验比较研究了分别以柘叶和桑叶Morus alba饲养家蚕后其消化液中抗病毒蛋白活性的差异。结果表明: 家蚕经柘叶饲养后消化液中红色荧光蛋白(red fluorescent protein, RFP)的强度无明显变化, 但柘叶饲养蚕消化液中脂肪酶、 胰蛋白酶的活性显著低于桑叶饲养蚕, 柘叶饲养蚕消化液中脂肪酶和胰蛋白酶的活性分别为1 421.71±202.60 U/L和19.67±8.17 U/mL, 桑叶饲养蚕脂肪酶和胰蛋白酶的活性分别为1 976.03±139.92 U/L和199.18±181.71 U/mL。这些结果说明, 消化液中脂肪酶和胰蛋白酶的活性水平低与柘叶饲养蚕易感染核型多角体病毒(BmNPV)可能相关。 相似文献
45.
Mussaenda yunnanensis, a new dioecious species of Rubiaceae from Yunnan Province, China, is described and illustrated. The new species can be recognized by its slender stem, congested‐cymose inflorescences and long corolla tubes. Differences between M. yunnanensis and two morphologically similar species (M. pubescens and M. antiloga) are presented. We also provide a key to all dioecious species of Mussaenda in China. The delimitation of the new species is further supported by molecular phylogenetic analyses based on eight plastid loci. 相似文献
46.
p53-dependent down-regulation of telomerase is mediated by p21waf1 总被引:13,自引:0,他引:13
Shats I Milyavsky M Tang X Stambolsky P Erez N Brosh R Kogan I Braunstein I Tzukerman M Ginsberg D Rotter V 《The Journal of biological chemistry》2004,279(49):50976-50985
47.
Formation of Adeno-Associated Virus Circular Genomes Is Differentially Regulated by Adenovirus E4 ORF6 and E2a Gene Expression 总被引:2,自引:7,他引:2 下载免费PDF全文
Dongsheng Duan Prerna Sharma Lorita Dudus Yulong Zhang Salih Sanlioglu Ziying Yan Yongping Yue Yihong Ye Rachael Lester Jusan Yang Krishna J. Fisher John F. Engelhardt 《Journal of virology》1999,73(1):161-169
A central feature of the adeno-associated virus (AAV) latent life cycle is persistence in the form of both integrated and episomal genomes. However, the molecular processes associated with episomal long-term persistence of AAV genomes are only poorly understood. To investigate these mechanisms, we have utilized a recombinant AAV (rAAV) shuttle vector to identify circular AAV intermediates from transduced HeLa cells and primary fibroblasts. The unique structural features exhibited by these transduction intermediates included circularized monomer and dimer virus genomes in a head-to-tail array, with associated specific base pair alterations in the 5′ viral D sequence. In HeLa cells, the abundance and stability of AAV circular intermediates were augmented by adenovirus expressing the E2a gene product. In the absence of E2a, adenovirus expressing the E4 open reading frame 6 gene product decreased the abundance of AAV circular intermediates, favoring instead the linear replication form monomer (Rfm) and dimer (Rfd) structures. In summary, the formation of AAV circular intermediates appears to represent a new pathway for AAV genome conversion, which is consistent with the head-to-tail concatemerization associated with latent-phase persistence of rAAV. A better understanding of this pathway may increase the utility of rAAV vectors for gene therapy. 相似文献
48.
Dynamic distribution of epidermal growth factor during mouse embryo peri-implantation 总被引:6,自引:0,他引:6
Embryo implantation depends on the synchronized development of the blastocyst and the endometrium. This process is highly controlled by the coordinated action of the steroid hormones: estrogen and progesterone. By autocrine, paracrine or juxtacrine routes, some growth factors or cytokines are involved in this steroidal regulation pathway. Here we report the effects of epidermal growth factor (EGF) on embryo implantation in the mouse, the expression and distribution patterns of EGF protein in the mouse blastocyst, ectoplacental cone (EPC) and peri-implantation uterus on days 1-8 of gestation.By RT-PCR and dot blot, we found that EGF and its receptor (EGFR) are co-expressed in the blastocyst and peri-implantational uteri of pregnant days 2-8 (D2-D8) mice. Injection of EGF antibody into a uterine horn on the third day of pregnancy (D3) significantly reduced the number of mouse embryos that implanted on D8, indicating EGF have a function in the mouse embryo implantation.Further investigation by using indirect immunofluorescence and confocal microscope was made to trace EGF and EGFR protein localization during the mouse embryo implantation. EGF and EGFR are co-localized in the blastocyst, and in the secondary trophoblastic giant cells (SGC) of the EPC. At the pre-implantation stage, the distribution of EGF protein in the mouse uterus changes from epithelium to stroma. On D1 of pregnancy, EGF is mainly distributed in uterine stroma and myometrium. On D2, it is present in the uterine epithelium. On D3, it changes again from the uterine epithelium to the stroma. By D4, EGF is predominantly in the stroma. This dynamic distribution correlates with the proliferation activity of uterine cells at each period. On D6-D8 of embryo implantation, EGF 3 protein accumulates at the uterine mesometrial pole, a region that contributes to the trophoblastic invasiveness and placentation.This temporal and spatial localization of EGF protein in the mouse uterus implicates the cytokine in the regulation of trophoblastic invasiveness and uterine receptiveness. 相似文献
49.
目的了解麻疹减毒活疫苗检定用Vero细胞代次的稳定性范围。方法用DMEM培养液将Vero细胞137代连续传至173代,检测其细胞活力,并观察Vero细胞的形态,接种麻疹减毒活疫苗参考品进行病毒滴定及热稳定性试验。结果 170代以下Vero细胞形态良好,细胞活力在85%以上,麻疹减毒活疫苗病毒滴度热稳定性试验的结果:137~169代Vero细胞活力及原麻疹减毒活疫苗病毒滴度热稳定性试验无统计学差异(P>0.05),170代以上代次的Vero细胞活力及原麻疹减毒活疫苗热稳定性试验有明显统计学差异(P<0.01)。结论 169代以下Vero细胞为检定用麻疹减毒活疫苗最佳细胞代次。 相似文献
50.