全文获取类型
收费全文 | 4720篇 |
免费 | 236篇 |
国内免费 | 409篇 |
出版年
2023年 | 50篇 |
2022年 | 75篇 |
2021年 | 100篇 |
2020年 | 96篇 |
2019年 | 130篇 |
2018年 | 131篇 |
2017年 | 93篇 |
2016年 | 101篇 |
2015年 | 136篇 |
2014年 | 280篇 |
2013年 | 287篇 |
2012年 | 231篇 |
2011年 | 302篇 |
2010年 | 221篇 |
2009年 | 238篇 |
2008年 | 255篇 |
2007年 | 321篇 |
2006年 | 230篇 |
2005年 | 187篇 |
2004年 | 146篇 |
2003年 | 135篇 |
2002年 | 118篇 |
2001年 | 102篇 |
2000年 | 86篇 |
1999年 | 107篇 |
1998年 | 72篇 |
1997年 | 71篇 |
1996年 | 67篇 |
1995年 | 68篇 |
1994年 | 54篇 |
1993年 | 62篇 |
1992年 | 65篇 |
1991年 | 64篇 |
1990年 | 48篇 |
1989年 | 37篇 |
1988年 | 32篇 |
1987年 | 49篇 |
1986年 | 24篇 |
1985年 | 63篇 |
1984年 | 53篇 |
1983年 | 48篇 |
1982年 | 39篇 |
1981年 | 37篇 |
1980年 | 49篇 |
1979年 | 33篇 |
1978年 | 37篇 |
1977年 | 25篇 |
1976年 | 29篇 |
1975年 | 25篇 |
1973年 | 21篇 |
排序方式: 共有5365条查询结果,搜索用时 125 毫秒
61.
Microspore culture is a very important and useful tool in plant breeding for haploid production and has been developed for many years.Brassica campestris (Brassica rapa L. ssp.oleifera) is an important oilseed crop, but it is relatively recalcitrant in tissue culture including microspore culture. The microspore culture in our laboratory is based on the Canadian protocol. Thirty genotypes ofB. campestris were included in this study; twenty produced embryos. The highest yield was 5930 embryos per 100 buds from Canadian genotype Cv-2, this result was one of the best that had been reported in microspore culture inB. campestris. The buds measuring 2.0 mm to 3.9 mm in length responded best to produce embryos, the optimum timing for microspore culture was confirmed to be during the mid-late to very-late uninucleate stage. The buds could be removed from either the main raceme or lateral racemes. Activated charcoal (150 mg l-1) was added to the liquid NLN medium, it promoted embryogenesis significantly; embryo development was faster and the embryo yield was significantly higher than those cultures without activated charcoal. The donor plant condition was considered an important factor influencing embryogenesis; older donor plants (older than five weeks) and a cold treatment are recommended. 相似文献
62.
We designed an Integrated Media Preparation System (IMPS) for continuous, on-line preparation of cell culture media and delivery to intermediate storage vessels or directly to a bioreactor. Key components of the IMPS include: a high precision, continuous fluid mixing device; formulation-specific liquid medium concentrates; validated process controls and membrane filtration; and automated dispensing into large volume flexible plastic containers. The IMPS system is designed to produce sterile, single-strength liquid medium from common raw materials at a delivery rate of 1000–3000 liters per hour and will manufacture homogenous batches from several thousand liters to over 60,000 liters. Fortified nutrient media prepared from multi-component 50X concentrates have been demonstrated to accelerate bioreactor seed chains, increase product yield, and reduce the overall manufacturing cost of nutrient medium. A productivity matrix will analyze the fully-loaded costs and contrast alternative methods for media preparation against projected biological yield.Abbreviations IMPS
Integrated Media Preparation System
- 50X
Nutrient fluid components formulated at fifty-fold final use concentration
- 1X
Nutrient fluid formulated at final, single-strength use concentration
- cGMP
Current Good Manufacturing Practices
- SCADA
Supervisory Control and Data Acquisition
- PLC
Process Logic Controller
- LTI
Life Technologies, Inc.
- WFI
Water for Injection
- CIP
Clean in place
- SIP
Sterilize in place
- HPLC
High performance liquid chromatography
- DMEM
Dulbecco's Modified Eagle's Medium 相似文献
63.
Hybridomas were cultured under steady-state conditions in a series of two continuous stirred-tank reactors (CSTRs), using a serum-free medium. The substrate not completely converted in the first CSTR, was transported with the cells to the second one and very low growth rates, high death rates, and lysis of viable cells were observed in this second CSTR. These conditions are hardly accessible in a single vessel, because such experiments would be extremely time-consuming and unstable due to a low viability. In contrast to what is often observed in literature, kinetic parameters could thus be derived without the neccessity for extrapolation to lower growth rates. Good agreement with literature averages for other hybridomas was found. Furthermore, showing that the reactor series is a valuable research tool for kinetic studies under extreme conditions, the possibility to observe cell death under stable and defined steady-state conditions offers interesting opportunities to investigate apoptosis and necrosis. Additionally, a model was developed that describes hybridoma growth and monoclonal antibody production in the bioreactor cascade on the basis of glutamine metabolism. Good agreement between the model and the experiments was found.Abbreviation MAb
Monoclonal antibody
Nomenclature
C
AConcentration of any (mol m-3) component A
D Dilution rate (s-1)
K
dDeath-rate constant (mol m-3)
K
lLysis-rate constant (mol m-3)
K
sMonod constant (mol m-3)
m Maintenance coefficient (mol cell-1 s-1)
q Specific consumption (mol cell-1 s-1) or production rate
t Time (s)
X Cell concentration (cell m-3)
Y Yield coefficient (cell mol-1)
Greek symbols
d
Specific death rate (s-1)
l
Specific lysis rate (s-1) of viable cells
net
Net specific growth (s-1) rate
true
True specific growth (s-1) rate 相似文献
64.
We have established a primary culture system for Drosophila eye imaginal discs. With this system, we were able to obtain neurite outgrowth from intact eye discs, eye disc fragments, and dissociated eye imaginal disc cells. Immunoreactivity to antibody 24B10 indicates that these extending neurites are photoreceptor axons. Three culture media were tested for their ability to support the survival of and neurite extension from eye disc fragments in vitro at 23°C. These, with supplements, were: five parts of Schneider's Drosophila medium with four parts of basal Eagle's medium (“4+5”); Leibovitz's L-15 medium (L-15); and Shields and Sang's M3 modified medium (MM3). We obtained the best results with MM3 supplemented with 2% fetal bovine serum (FBS). Eye disc fragments survived in this medium for at least 20 days. Pigmentation in the nonphotoreceptor pigment cells in cultures from the prepupa required the presence of 20-hydroxyecdysone (20-HE) (1 μg/ml), whereas neurite outgrowth was seen in the absence of 20-HE. Donor animals had to fall within a range of ages to obtain appropriate eye disc differentiation in vitro. Eye discs from 5-h pupae (P+5) or older commenced ommachrome synthesis in vitro, in a temporal sequence close to that found in vivo, whereas the in vitro synthesis of this pigment was delayed in eye discs from younger flies. Average neurite length was not affected by age among pupae younger than P+5; but neurite outgrowth from P+24 was scarce, probably because by this time photoreceptor axons had already grown in vivo and were severed and unable to regenerate in vitro. Eye discs taken from third instar larvae or white prepupae continued their mitotic activity in vitro. Together with the advance of the morphogenetic furrow at the leading edge of retinal development, this observation is consistent with the evidence that pattern formation continues in vitro. Morphogenetic changes were manifested in cultures. Viability tests with calcein AM and ethidium bromide revealed few dead cells in living cultures. © 1995 John Wiley & Sons, Inc. 相似文献
65.
本文报告一种为适于杂交瘤细胞生长“无血清”培养基-适量的成牛血清低分子成分超滤液、10mg/L转铁蛋白(T)、10mg/L胰岛素(I)、20μmol/L乙醇胺(E)、40nmol/L硒酸钠(S)等诸补充成分替代胎牛血清加到基础液中-也完全适用于培养肿瘤细胞,且观察到与之相似的规律性结果,癌细胞在本“无血清”培养若的生长水平达到在含胎牛血清培养基中生长的水平。对于癌细胞的生长,LMW-CS与T、I、 相似文献
66.
屎克螂适宜的繁殖条件初步研究 总被引:2,自引:0,他引:2
1991—1994年对屎克螂适宜的繁殖条件研究结果:1.土层40cm和60cm的平均温度分别为15.92±6.34℃和15.89±6.08℃;平均土壤含水量分别为26.70±3.74%和23.72±1.88%,使各虫态处于一个几乎恒温恒湿的条件,有利于生长发育与越冬,2.土壤含水量17%、20%、22%、26%和29%,其产卵最多的深度分别在65─70cm,75─80cm,65─80cm,65—80cm和35—50cm,分别占产卵总数的34.16%、42.42%、68.42%、55.40%和90.56%。3.不同土壤的产卵深度,沙壤土以土下45—50cm处最多,占产卵总数的34.78%;沙土以65—85cm最多,占产卵总数的86.67%;粘土以45—60cm最多,占产卵总数的51.11%。 相似文献
67.
在自生异养条件下,旋扭山绿豆根瘤菌MXDI6菌株的氢酶诱导表达受气相、pH值、镍等因子影响:氢酶表达的最适氧浓度为4%,最适氢浓度为15%,二氧化碳没有明显影响;氢酶表达的pH值以5.0—6.0为宜;0.5μmol/LNiCl2明显促进吸氢活性,但镍浓度大于1μmol/L则抑制吸氢活性. 相似文献
68.
Production of Serine Proteases by the Oyster Pathogen Perkinsus marinus (Apicomplexa) In Vitro 总被引:1,自引:1,他引:0
JEROME F. LA PEYRE DORIS Y. SCHAFHAUSER ESAM H. RIZKALLA MOHAMED FAISAL 《The Journal of eukaryotic microbiology》1995,42(5):544-551
ABSTRACT. Analysis of the cell-free supernatants of Perkinsus marinus cultures by sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and silver staining revealed the presence of as many as 17 bands ranging in molecular weight from 239 to 32 kDa. These bands were not present in un-inoculated medium. Moreover, P. marinus produces extracellular proteins that possess proteolytic activities; the cell-free supernatants of P. marinus cultures could digest a variety of proteins including gelatin, casein, fibronectin and laminin. Oyster plasma was also digested by cell-free culture supernatants. The proteolytic activity in cell-free culture supernatants was detected 24 h post-inoculation, while no proteolytic activity could be detected in cell lysates. The proteolytic activities were characterized using substrate-impregnated sodium dodecylsulfate-polyacrylamide gels and had approximate molecular weights ranging from 55 to 35 kDa. The proteolytic activity of cell-free culture supernatants was inhibited by the serine protease inhibitors phenylmethylsulphonyl fluoride, 3,4-dichloroisocoumarin and soybean trypsin inhibitor. In contrast, inhibitors (i.e. trans-epoxysuccinyll-leucylamido(4-guanidino)-butane, 1, 10-phenanthroline, captopril, ethylenediaminetetracetic acid, pepstatin A or diazoacetyl-DL-norleucine methyl ester) from the other three classes of proteases had no effect. It was concluded that the P. marinus proteases in cell-free culture supernatants are serine proteases. 相似文献
69.
Koichiro Yoshihara Motokatsu Tsuyuki Asako Itaya Yasuharu Tanaka Tomoya Kamiya 《Molecular and cellular biochemistry》1994,135(2):143-151
HL-60 cells transferred from serum-supplemented to serum-free culture medium initially bound to culture plate tightly and then released from the plate on increasing the culture time and resumed exponential growth after about 8 h lag. At the initial stage of the culture, the cells became extremely sensitive to 3-aminobenzamide, a potent inhibitor of poly (ADP-ribose) polymerase, and, at 1 mM, 80 to 90% of the cells were lysed within 20 h, whereas the inhibitor was totally ineffective on the cell growth in serum-supplemented medium at the concentration. Non-inhibitory analogs of the inhibitor were ineffective. Assay of poly(ADP-ribose) polymerase activity in permeable cells indicated that a transient activation of the enzyme occurred during the culture in serum-free medium (the maximum activation was observed at 8 h of the culture). The cells conditioned in serum-free medium for 24 h acquired significant resistancy to the inhibitor. A low concentration of fibronectin (5 to 10/ml) and a relatively high concentration of bovine serum albumin (0.5 to 1 mg/ml) effectively blocked the cell attachment to plate and also the 3-aminobenzamide-induced cell lysis. These results suggest that poly(ADP-ribose) polymerase is involved in a process essential for HL-60 cells to adapt to a serumdeprived growth condition. 相似文献
70.
水平回转对水稻幼苗叶细胞的影响 总被引:9,自引:0,他引:9
对在模拟微重力装置上回转14 天的水稻幼苗叶细胞进行了亚显微形态、电子探针和细胞酶化学研究。发现叶细胞质膜上Ca2+ -ATP酶活性消失,膜内钙总量上升、膜外钙总量下降,细胞骨架变得疏松,细胞壁变薄并凹凸不平。叶绿体的基粒和线粒体的内嵴亦有部分变化。其变化机制,首先是细胞质膜上Ca2+ -ATP酶活性消失,膜上钙泵停止工作,跨膜钙浓度差减小,膜内钙浓度上升,微管、微丝聚合受阻,细胞骨架疏松,分泌泡移动失去导向,从而导致细胞壁变薄等状态 相似文献