全文获取类型
收费全文 | 10212篇 |
免费 | 908篇 |
国内免费 | 892篇 |
专业分类
12012篇 |
出版年
2024年 | 29篇 |
2023年 | 161篇 |
2022年 | 356篇 |
2021年 | 599篇 |
2020年 | 422篇 |
2019年 | 454篇 |
2018年 | 450篇 |
2017年 | 323篇 |
2016年 | 456篇 |
2015年 | 639篇 |
2014年 | 841篇 |
2013年 | 802篇 |
2012年 | 953篇 |
2011年 | 803篇 |
2010年 | 554篇 |
2009年 | 441篇 |
2008年 | 481篇 |
2007年 | 503篇 |
2006年 | 421篇 |
2005年 | 345篇 |
2004年 | 304篇 |
2003年 | 234篇 |
2002年 | 203篇 |
2001年 | 175篇 |
2000年 | 156篇 |
1999年 | 154篇 |
1998年 | 71篇 |
1997年 | 69篇 |
1996年 | 74篇 |
1995年 | 53篇 |
1994年 | 64篇 |
1993年 | 51篇 |
1992年 | 75篇 |
1991年 | 59篇 |
1990年 | 39篇 |
1989年 | 51篇 |
1988年 | 29篇 |
1987年 | 33篇 |
1986年 | 24篇 |
1985年 | 25篇 |
1984年 | 8篇 |
1983年 | 11篇 |
1982年 | 3篇 |
1981年 | 1篇 |
1980年 | 5篇 |
1979年 | 6篇 |
1978年 | 1篇 |
1970年 | 1篇 |
排序方式: 共有10000条查询结果,搜索用时 15 毫秒
81.
82.
Changlin Fu William P. Donovan Olga Shikapwashya-Hasser Xudong Ye Robert H. Cole 《PloS one》2014,9(12)
Molecular cloning is utilized in nearly every facet of biological and medical research. We have developed a method, termed Hot Fusion, to efficiently clone one or multiple DNA fragments into plasmid vectors without the use of ligase. The method is directional, produces seamless junctions and is not dependent on the availability of restriction sites for inserts. Fragments are assembled based on shared homology regions of 17–30 bp at the junctions, which greatly simplifies the construct design. Hot Fusion is carried out in a one-step, single tube reaction at 50°C for one hour followed by cooling to room temperature. In addition to its utility for multi-fragment assembly Hot Fusion provides a highly efficient method for cloning DNA fragments containing inverted repeats for applications such as RNAi. The overall cloning efficiency is in the order of 90–95%. 相似文献
83.
耦合中空纤维膜超滤分离游离细胞催化合成ATP 总被引:1,自引:0,他引:1
对耦合中空纤维膜超滤分离进行游离细胞催化合成ATP过程进行了实验研究,考察了细胞的催化效率和膜组件的操作稳定性。结果显示,中空纤维超滤膜的耦合分离能有效地截留反应液中的游离酶,其中乙醇脱氢酶(ADH)和已糖激酶(HK)的稳态截留效率在95%以上。耦合膜分离的酵母细胞催化ATP合成反应可重复使用2.5~3.0次,酶的利用率比普通分离的细胞提高2.0~2.5倍。中空纤维超滤膜于0.1Mpa工作压力下连续11批耦合分离操作,膜的渗透性无明显下降,过滤速率保持在初速率的95%以上。在稀释速率0.25h-1下,反应体系保持了连续5h的ATP高转化率合成与分离耦合的拟稳态操作。 相似文献
84.
85.
86.
87.
Inositol pyrophosphates are required for DNA hyperrecombination in protein kinase c1 mutant yeast 总被引:5,自引:0,他引:5
Diphosphoinositol pentakisphosphate (InsP(7)) and bis-diphosphoinositol tetrakisphosphate (InsP(8)) contain energetic pyrophosphate groups, occur throughout animal and plant kingdoms, and are synthesized by a recently cloned family of inositol hexakisphosphate kinases (InsP(6)Ks). We report that these inositol pyrophosphates mediate homologous DNA recombination in yeast S. cerevisae. Hyperrecombination, caused by altered protein kinase C1 (PKC1), is lost in yeast with deletion of yeast InsP(6)K (yInsP(6)K) and can be restored selectively by catalytically active yeast or mammalian InsP(6)Ks. Inositol pyrophosphates are required for two forms of hyperrecombination that differ in mechanism, suggesting some generalities for actions of inositol pyrophosphates in recombination. 相似文献
88.
Hydrolysis of lignocellulosic materials for ethanol production: a review 总被引:85,自引:0,他引:85
Lignocellulosic biomass can be utilized to produce ethanol, a promising alternative energy source for the limited crude oil. There are mainly two processes involved in the conversion: hydrolysis of cellulose in the lignocellulosic biomass to produce reducing sugars, and fermentation of the sugars to ethanol. The cost of ethanol production from lignocellulosic materials is relatively high based on current technologies, and the main challenges are the low yield and high cost of the hydrolysis process. Considerable research efforts have been made to improve the hydrolysis of lignocellulosic materials. Pretreatment of lignocellulosic materials to remove lignin and hemicellulose can significantly enhance the hydrolysis of cellulose. Optimization of the cellulase enzymes and the enzyme loading can also improve the hydrolysis. Simultaneous saccharification and fermentation effectively removes glucose, which is an inhibitor to cellulase activity, thus increasing the yield and rate of cellulose hydrolysis. 相似文献
89.
Jun Zhao Shaoyi Wang Jianqiang Bao Xiaojuan Sun Xiaochen Zhang Xiuli Zhang Dongxia Ye Jie Wei Changsheng Liu Xinquan Jiang Gang Shen Zhiyuan Zhang 《PloS one》2013,8(1)
Calcium phosphate (Ca-P) scaffolds have been widely employed as a supportive matrix and delivery system for bone tissue engineering. Previous studies using osteoinductive growth factors loaded Ca-P scaffolds via passive adsorption often experience issues associated with easy inactivation and uncontrolled release. In present study, a new delivery system was fabricated using bone morphogenetic protein-2 (BMP-2) loaded calcium-deficient hydroxyapatite (CDHA) scaffold by lyophilization with addition of trehalose. The in vitro osteogenesis effects of this formulation were compared with lyophilized BMP-2/CDHA construct without trehalose and absorbed BMP-2/CDHA constructs with or without trehalose. The release characteristics and alkaline phosphatase (ALP) activity analyses showed that addition of trehalose could sufficiently protect BMP-2 bioactivity during lyophilization and achieve sustained BMP-2 release from lyophilized CDHA construct in vitro and in vivo. However, absorbed BMP-2/CDHA constructs with or without trehalose showed similar BMP-2 bioactivity and presented a burst release. Quantitative real-time PCR (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA) demonstrated that lyophilized BMP-2/CDHA construct with trehalose (lyo-tre-BMP-2) promoted osteogenic differentiation of bone marrow stromal cells (bMSCs) significantly and this formulation could preserve over 70% protein bioactivity after 5 weeks storage at 25°C. Micro-computed tomography, histological and fluorescent labeling analyses further demonstrated that lyo-tre-BMP-2 formulation combined with bMSCs led to the most percentage of new bone volume (38.79% ±5.32%) and area (40.71% ±7.14%) as well as the most percentage of fluorochrome stained bone area (alizarin red S: 2.64% ±0.44%, calcein: 6.08% ±1.37%) and mineral apposition rate (4.13±0.62 µm/day) in critical-sized rat cranial defects healing. Biomechanical tests also indicated the maximum stiffness (118.17±15.02 Mpa) and load of fracture (144.67±16.13 N). These results lay a potential framework for future study by using trehalose to preserve growth factor bioactivity and optimize release profile of Ca-P based delivery system for enhanced bone regeneration. 相似文献
90.
Huijuan Liu Jia Ru Jiuhui Qu Ruihua Dai Zijian Wang Chun Hu 《Bioresource technology》2009,100(12):2995-3002
A new biomimetic absorbent, cellulose acetate (CA) embedded with triolein (CA-triolein), was prepared and applied for the removal of persistent organic pollutants (POPs) from micro-polluted aqueous solution. The comparison of CA-triolein, CA and granular activated carbon (GAC) for dieldrin removal was investigated. Results showed that CA-triolein absorbent gave a lowest residual concentration after 24 h although GAC had high removal rate in the first 4 h adsorption. Then the removal efficiency of mixed POPs (e.g. aldrin, dieldrin, endrin and heptachlor epoxide), absorption isotherm, absorbent regeneration and initial column experiments of CA-triolein were studied in detail. The linear absorption isotherm and the independent absorption in binary isotherm indicated that the selected POPs are mainly absorbed onto CA-triolein absorbent by a partition mechanism. The absorption constant, K, was closely related to the hydrophobic property of the compound. Thermodynamic calculations showed that the absorption was spontaneous, with a high affinity and the absorption was an endothermic reaction. Rinsing with hexane the CA-triolein absorbent can be regenerated after absorption of POPs. No significant decrease in the dieldrin removal efficiency was observed even when the absorption–regeneration process was repeated for five times. The results of initial column experiments showed that the CA-triolein absorbent did not reach the breakthrough point at a breakthrough empty-bed volume (BV) of 3200 when the influent concentration was 1–1.5 μg/L and the empty-bed contact time (EBCT) was 20 min. 相似文献