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101.
Plant regeneration from mesophyll protoplasts of Agrobacterium rhizogenes-transformed Astragalus melilotoides 总被引:1,自引:1,他引:0
Plant regeneration from mesophyll protoplasts of Agrobacterium rhizogenes-transformed Astragalus melilotoides Pall. was here developed. The protoplasts were isolated directly from the leaves of the hairy root-induced plants. The highest
yield of protoplasts was obtained from fully expanded leaves of young plants. Their viability was up to 72 ± 2.3 %. The highest
division frequency (32.4 ± 0.13 %) and sustained divisions were obtained in Durand, Potrykus and Donn (DPD) medium supplemented
with 2.0 mg dm−3 2,4-dichlorophenoxyacetic acid, 0.2 mg dm−3 6-benzylaminopurine, 0.3 M mannitol, 2 % sucrose and 500 mg dm−3 casein hydrolysate at the plating density of 3.0 × 105 cm−3. The frequency of shoot differentiation from protocalli reached to 91.75 ± 3.1 %. Opine synthesis and polymerase chain reaction
analysis confirmed that T-DNA still existed in the protoplast regenerated plants. 相似文献
102.
103.
Ca2+/H+ 反向转运体作为一类 Ca2+外向转运器,在植物的营养和信号转导中起着非常重要的作用 . 克隆了水稻 Ca2+/H+ 反向转运体基因 OsCAX3 ,序列分析表明 OsCAX3 具有 11 个跨膜区,其中在第 6 和第 7 个跨膜区之间有一个 17 个氨基酸组成的酸性基序 (acid motif) ,功能互补实验证明 OsCAX3 具有转运 Ca2+ 的功能,并且其 N 端 26 个氨基酸序列对转运 Ca2+ 具有一定的抑制作用 . RT-PCR 分析表明 OsCAX3 的表达受到外源 Ca2+ 的诱导 . 利用 PSORT prediction 进行亚细胞定位分析,和利用 OsCAX3-GFP 融合蛋白瞬时表达分析证明, OsCAX3 定位于细胞质膜 . 以上结果表明, OsCAX3 是一种定位于细胞质膜上的 Ca2+/H+ 反向转运体 . 相似文献
104.
植物体细胞原生质体遗传转化研究 总被引:6,自引:1,他引:5
重点介绍了植物体细胞原生质体遗传转化的方法和当前已经取得的成果,同时提出了目前原生质体遗传转化中存在的问题,展望了今后的工作重点。植物原生质体遗传转化的方法主要有:PEG介导转化法、电击穿孔转化法、脂质体介导转化法、农杆菌共培养转化法等。 相似文献
105.
Giant protoplasts of Saccharomyces cerevisiae of 10-35 µm in diameter were generated by multi-cell electrofusion. Thereby two different preparation strategies were evaluated with a focus on size distribution and “patchability” of electrofused protoplasts. In general, parental protoplasts were suitable for electrofusion 1-12 h after isolation. The electrophysiological properties of electrofused giant protoplasts could be analyzed by the whole-cell patch clamp technique. The area-specific membrane capacitance (0.66 ± 0.07 µF/cm2) and conductance (23-44 µS/cm2) of giant protoplasts were consistent with the corresponding data for parental protoplasts. Measurements with fluorescein-filled patch pipettes allowed to exclude any internal compartmentalisation of giant protoplasts by plasma membranes, since uniform (diffusion-controlled) dye uptake was only observed in the whole-cell configuration, but not in the cell-attached formation. The homogeneous structure of giant protoplasts was further confirmed by the observation that no plasma membrane associated fluorescence was seen in the interior of giant cells after electrofusion of protoplasts expressing the light-activated cation channel Channelrhodopsin-2 (ChR2) linked to yellow fluorescent protein (YFP). Patch clamp analysis of the heterologously expressed ChR2-YFP showed typical blue light dependent, inwardly-directed currents for both electrofused giant and parental protoplasts. Most importantly, neither channel characteristics nor channel expression density was altered by electric field treatment. Summarising, multi-cell electrofusion increases considerably the absolute number of membrane proteins accessible in patch clamp experiments, thus presumably providing a convenient tool for the biophysical investigation of low-signal transporters and channels. 相似文献
106.
107.
The ratio of activities of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase (G6P DH/6PG DH), and the contents of glucose-6-phosphate (G6P), 6-phosphogluconate (6PG) and fructose-6-phosphate (F6P) were studied at various stages of potato virus Y (PVY) multiplication in Nicotiana tabacum cv. Samsun. G6P DH/6PG DH increased through the experiment from 0.42 to 0.53 in leaves of healthy tobacco, and up to 0.59 in PVY systemically infected leaves. However, these ratios in the ruptured protoplast preparations, and the chloroplast and cytosol fractions of healthy protoplasts were similar to that from infected ones. The ratio lower than 1, found in the healthy and/or PVY- infected leaf tissues and in the infected protoplasts as well, confirms the assumption that G6P DH is the control enzyme of oxidative pentosephosphate pathway not only in the healthy but also in the infected plants. The contents of G6P, 6PG and F6P in the period of the highest PVY multiplication were strongly decreased (to 30 – 50 % when compared with control healthy leaves) and were negatively correlated with the G6P DH and 6PG DH activities. 相似文献
108.
Joanna Augustynowicz Małgorzata Lekka Kveta Burda Halina Gabryś 《Acta Physiologiae Plantarum》2001,23(3):291-302
Translocations of chloroplasts induced by blue light were investigated in both leaves and protoplasts isolated from leaf mesophyll
of Nicotiana tabacum. In the leaf tissue, the responses of chloroplasts were similar to those observed in other, higher and lower plant species.
Weak and strong light induced movements of chloroplasts towards cell walls perpendicular and parallel to the light direction,
respectively. Treatment with cytochalasin D, an actin-disturbing agent, blocked the movements. This shows that actin is involved
in the motile system of chloroplast translocation in tobacco. By monitoring the response of chloroplasts to light in isolated
protoplasts, we addressed the question whether the presence of the cell wall is necessary for the translocations of chloroplasts
to occur. In control protoplasts (isolated at room temperature from unstressed leaves), no clear light intensity-dependent
changes were observed in chloroplast distribution pattern. In contrast, in protoplasts obtained from plants treated with 4
°C for 8 h the chloroplasts maintained their responsiveness to light. Atomic Force Microscopy was used to measure elastic
properties of the protoplasts. Young’s modulus, which reflects rigidity of the material, was 10 times higher for protoplasts
of the coldstressed plants as compared to those isolated from the control plants. The rigidity of protoplasts isolated from
the plants treated with low temperature was reduced four-fold by exposure to cytochalasin D. It appears that the status of
protoplast actin is a factor responsible for elasticity of protoplasts. We speculate that unknown, cold stress-induced factors,
maintain the orientational movements due to anchorage of the actin cytoskeleton in the plasma membrane despite the cell wall
removal. 相似文献
109.
The construction of inulin-assimilating and sorbitol-producing fusants was achieved by intergeneric protoplast fusion between Kluyveromyces sp. Y-85 and Saccharomyces cerevisiae E-15. The cells of parental strains were pretreated with 0.1% EDTA (w/v) and 2-mercaptoethanol (0.1%, v/v) and then exposed to 2.0% (w/v) Zymolase at 30 °C for 30–40 min. The optimized fusion condition demonstrated that with the presence of 30% (w/v) polyethylene glycol 6000 (PEG-6000) and 10 mM CaCl2 for 30 min, the fusion frequency reached 2.64 fusants/106 parental cells. The fusants were screened by different characters between two parental strains and further identified by DNA contents, inulinase activity and sorbitol productivity. One of the genetically stable fusants, Strain F27, reached a maximal sorbitol production of 4.87 g/100 ml under optimal fermentation condition. 相似文献
110.