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141.
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We previously reported that substitution of Arg258 within the switch 3 region of Gsalpha impaired activation and increased basal GDP release due to loss of an interaction between the helical and GTPase domains (Warner, D. R., Weng, G., Yu, S., Matalon, R., and Weinstein, L. S. (1998) J Biol. Chem. 273, 23976-23983). The adjacent residue (Glu259) is strictly conserved in G protein alpha-subunits and is predicted to be important in activation. To determine the importance of Glu259, this residue was mutated to Ala (Gsalpha-E259A), Gln (Gsalpha-E259Q), Asp (Gsalpha-E259D), or Val (Gsalpha-E259V), and the properties of in vitro translation products were examined. The Gsalpha-E259V was studied because this mutation was identified in a patient with Albright hereditary osteodystrophy. S49 cyc reconstitution assays demonstrated that Gsalpha-E259D stimulated adenylyl cyclase normally in the presence of GTPgammaS but was less efficient with isoproterenol or AlF4-. The other mutants had more severely impaired effector activation, particularly in response to AlF4-. In trypsin protection assays, GTPgammaS was a more effective activator than AlF4- for all mutants, with Gsalpha-E259D being the least severely impaired. For Gsalpha-E259D, the AlF4--induced activation defect was more pronounced at low Mg2+ concentrations. Gsalpha-E259D and Gsalpha-E259A purified from Escherichia coli had normal rates of GDP release (as assessed by the rate GTPgammaS binding). However, for both mutants, the ability of AlF4- to decrease the rate of GTPgammaS binding was impaired, suggesting that they bound AlF4- more poorly. GTPgammaS bound to purified Gsalpha-E259D irreversibly in the presence of 1 mM free Mg2+, but dissociated readily at micromolar concentrations. Sucrose density gradient analysis of in vitro translates demonstrated that all mutants except Gsalpha-E259V bind to beta gamma at 0 degreesC and were stable at higher temperatures. In the active conformation Glu259 interacts with conserved residues in the switch 2 region that are important in maintaining both the active state and AlF4- in the guanine nucleotide binding pocket. Although both Gsalpha Arg258 and Glu259 are critical for activation, the mechanisms by which these residues affect Gsalpha protein activation are distinct. 相似文献
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H Tanaka CA Desouza PP Jones ET Stevenson KP Davy DR Seals 《Canadian Metallurgical Quarterly》1997,83(6):1947-1953
Using a meta-analytic approach, we recently reported that the rate of decline in maximal oxygen uptake (VO2 max) with age in healthy women is greatest in the most physically active and smallest in the least active when expressed in milliliters per kilogram per minute per decade. We tested this hypothesis prospectively under well-controlled laboratory conditions by studying 156 healthy, nonobese women (age 20-75 yr): 84 endurance-trained runners (ET) and 72 sedentary subjects (S). ET were matched across the age range for age-adjusted 10-km running performance. Body mass was positively related with age in S but not in ET. Fat-free mass was not different with age in ET or S. Maximal respiratory exchange ratio and rating of perceived exertion were similar across age in ET and S, suggesting equivalent voluntary maximal efforts. There was a significant but modest decline in running mileage, frequency, and speed with advancing age in ET. VO2 max (ml . kg-1 . min-1) was inversely related to age (P < 0.001) in ET (r = -0.82) and S (r = -0.71) and was higher at any age in ET. Consistent with our meta-analysic findings, the absolute rate of decline in VO2 max was greater in ET (-5.7 ml . kg-1 . min-1 . decade-1) compared with S (-3.2 ml . kg-1 . min-1 . decade-1; P < 0. 01), but the relative (%) rate of decline was similar (-9.7 vs -9. 1%/decade; not significant). The greater absolute rate of decline in VO2 max in ET compared with S was not associated with a greater rate of decline in maximal heart rate (-5.6 vs. -6.2 beats . min-1 . decade-1), nor was it related to training factors. The present cross-sectional findings provide additional evidence that the absolute, but not the relative, rate of decline in maximal aerobic capacity with age may be greater in highly physically active women compared with their sedentary healthy peers. This difference does not appear to be related to age-associated changes in maximal heart rate, body composition, or training factors. 相似文献
145.
GS Firestein F Echeverri M Yeo NJ Zvaifler DR Green 《Canadian Metallurgical Quarterly》1997,94(20):10895-10900
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Follicles > or = 5 mm were ablated in pony mares by a transvaginal ultrasound-guided technique on Day 10 (ovulation = Day 0). Follicle emergence (at 15 mm, experiment 1; at 6 mm, experiment 2) and development of the new wave was monitored by transrectal ultrasound. Deviation was defined as the beginning of a marked difference in growth rates between the two largest follicles. In experiment 1, mares were grouped (n = 4 per group) into controls, ablation-controls (ablations at Day 10 only), and a two-follicle model (periodic ablation sessions so that only the two largest follicles developed). There were no significant indications that the two-follicle model altered follicle diameters, growth rates, or time intervals of the two retained follicles at or between events (follicle emergence, deviation, and ovulation). In experiment 2, the two-follicle model (n = 14) was used for follicle and hormonal characterization and hypothesis testing, without the tedious and error-prone necessity for tracking many (e.g., 20) individual follicles. The future dominant follicle emerged a mean of 1 day earlier (p < 0.008) than the future subordinate follicle, the growth rates for the two follicles between emergence and deviation (6 days later) did not differ, and the dominant follicle was larger at the beginning of deviation (23.1 +/- 0.8 mm versus 19.6 +/- 0.9 mm; p < 0.0001). Mean FSH and LH concentrations increased (p < 0.05) concomitantly from emergence of the future dominant follicle and peaked 3 days later when the follicle was a mean of 13 mm. Thereafter, the two hormones disassociated until ovulation: FSH decreased and LH increased. Results supported the hypothesis that the future dominant follicle has an early size advantage over future subordinate follicles and indicated that the advantage was present as early as 6 days before deviation. 相似文献
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Prognostic classification for esophageal atresia and tracheoesophageal fistula: Waterston versus Montreal 总被引:1,自引:0,他引:1
S Teich DP Barton ME Ginn-Pease DR King 《Canadian Metallurgical Quarterly》1997,32(7):1075-9; discussion 1079-80
Since 1962, the Waterston classification has been used to stratify neonates who have esophageal atresia (EA) and/or tracheoesophageal fistula (TEF) into prognostic categories based on birth weight, the presence of pneumonia, and the identification of other congenital anomalies. In response to advances in neonatal care, the surgeons from the Montreal Children's Hospital proposed a new categorization system in 1993 in an attempt to define the current risk factors for patients who have EA/TEF. In the Montreal experience only two characteristics independently affected survival: preoperative ventilator dependence and associated major anomalies. The goal of this study was to determine which system had the greatest validity for the evaluation of prognosis in our patients with EA/TEF. The charts of 94 patients who had EA/TEF treated between 1972 and 1991 were reviewed. Patients were classified using both the Waterston and Montreal systems. Groups were compared with Fisher's Exact test using a 95% confidence level for statistical significance. Eleven infants were ventilator dependent preoperatively; 62 children had major associated anomalies, 8 of which were considered life threatening. Sixteen children died within 4 years, eight during their initial hospital stay. Five of the eight early postoperative deaths occurred in the highest-risk patients (Waterston C or Montreal II). Analysis was performed for multiple risk factors and mortality. As in the Montreal study, the presence of life-threatening and major congenital anomalies represented significant risk factors for death. Pulmonary disease as delineated by ventilator dependence appeared to be more accurate than pneumonia. This study confirms the accuracy of the Montreal classification in defining prognosis for EA/TEF. The Montreal system more accurately identifies children at highest risk than the Waterston classification. 相似文献