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41.
血管内皮生长因子受体启动子驱动重组腺病毒双自杀基因系统选择性杀伤大肠癌细胞 总被引:4,自引:0,他引:4
Rong-kai Lin Zong-hai Huang Guo-qiang Su Zhi-yong Ke Jian-xiong Chen Jin-jie Zhou 《第一军医大学学报》2005,25(5):524-527
OBJECTIVE: To observe the selective killing effect of adenovirus (Ad)-mediated double suicide gene driven by kinase domain-containing receptor(KDR) promoter on human colorectal cancer LoVo cells and human umbilical vein endothelial ECV304 cells. METHODS: The plasmid pAdEasy-KDR-CDglyTK was transfected into 293 packaging cells for amplification of the infectious Ad and used to infect the KDR-producing cells (ECV304 and LoVo) and the KDR-nonproducing cells (LS174T) respectively. The three cells were treated with the prodrugs 5-flurocytosine (5-FC) and ganciclovir (GCV) at different concentrations after infection. The killing effects of the fusion gene system on the cells were evaluated. The distribution of cell cycle was detected by flow cytometry. RESULTS: The infection rates of the recombinant Ad were similar among the 3 cells, gradually increasing with the increment of multiplicity of infection (MOI) and reaching 100% with the MOI of 200. The LoVo cells and ECV304 cells infected with Ad-KDR-CDglyTK were highly sensitive to both of the prodrugs (P>0.1), whereas the infected LS174T cells failed to exhibit similar sensitivity (P<0.001). The killing effect of CD/TK fusion gene on the target cells was much stronger than that of either suicide gene (P<0.001). The cell cycle of LoVo cells was arrested at G1 phase. CONCLUSION: The CD/TK fusion gene system driven by KDR promoter can selectively kill KDR-expressing human colorectal cancer LoVo cells and endothelial cells. 相似文献
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原代培养神经细胞膜经N—甲基—D—D天冬氨酸和MK—801作用后原子力微镜观察 总被引:4,自引:0,他引:4
目的:对比观察正常培养皮层神经元在N-甲基-D-天冬氨酸(NMDA)及其受体拮抗剂-MK-801作用下膜表面三维构像形态的改变。方法:利用分辨率为0.1-0.01nm的原子力显微镜(AFM)对原代培养大鼠皮层神经细胞膜表面进行纳米尺度的扫描观测。结果:正常神经元膜表面光滑,起伏均匀,隆起的颗粒状蛋白密集,间隔规律。NMDA损伤后神经元破碎,崩解,膜失去连续性,NMDA+MK-801作用下神经元膜皱折增加,边缘粗糙,起伏程度介于前两者之间,结论:(1)AFM具有分辨率高,制样简单特点,(2)AFM能细微地分辨损伤保护作用后引起的细胞膜表面三维形态改变。(3)NMDA作用后膜结构开始解体,膜蛋白颗粒聚集增大,脂质凹陷加深,间距增宽,表面粗糙度增加。 相似文献
44.
正常血钾型周期性麻痹SCN4A基因新突变的检测 总被引:7,自引:1,他引:6
目的 报道正常血钾型周期性麻痹 (normoPP)一家系的临床特点 ,并筛查SCN4A基因以期发现有义突变。方法 提取知情同意的患者及部分家属外周血基因组DNA ,应用变性高效液相色谱分析 (DHPLC)技术筛查患者SCN4A基因全部 2 4个外显子 ,对发现异常者进行测序分析。结果先证者常规实验室检查未见异常 ,发作期肌酸激酶 (CK) 112 6U/L(正常值 <2 0 0U/L) ,肌电图正常。发作间期行肌肉活检未见显著异常。基因研究发现先证者及其父亲 (患者 )SCN4A基因发生同一新突变G2 10 1A ,并引起氨基酸序列改变Arg6 75Gln。该突变不同于目前发现的明确导致高钾型周期性麻痹(hyperPP)的突变 ,也不同于已知的SCN4A基因所有突变。结论 normoPP患者存在一新突变Arg6 75Gln ,该突变可能与疾病相关。 相似文献
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46.
A two decade survey of respiratory adenovirus in Taiwan: the reemergence of adenovirus types 7 and 4 总被引:3,自引:0,他引:3
Lin KH Lin YC Chen HL Ke GM Chiang CJ Hwang KP Chu PY Lin JH Liu DP Chen HY 《Journal of medical virology》2004,73(2):274-279
From November 1999 to December 2001, three outbreaks of adenovirus (Ad) respiratory infection occurred in southern Taiwan. To determine the circulating serotypes and molecular epidemiology, a total of 524 virus strains were randomly selected from 1,064 strains isolated from 1981 to 2001, and were studied using restriction fragment length polymorphism (RFLP) and polymerase chain reaction (PCR)-RFLP. The major subgenus found was subgenus B (45%), followed by subgenus E (29%) and subgenus C (25%). Ad3 and Ad7 were the major types found during the 1st outbreak, which occurred from November 1999 to March 2000, while Ad4 was found mainly during the 2nd and 3rd outbreaks in October 2000 and September 2001, respectively. Both Ad7 and Ad4 were reemerged serotypes, whereas Ad3 was consistently isolated during the survey, although it declined drastically from 36 to 2% in 2001. Genotype analysis in this study showed that the only strain of Ad7 found in 1983 was Ad7a, but all randomly selected strains of Ad7 isolated during 1999-2000 were Ad7b. The clinical features of 217 patients were analyzed during the 1999-2000 outbreaks. About 79% of the total cases were less than 7 years old. The ratio of male to female was 2:1. Severe infections, such as pneumonia and acute bronchitis, accounted for nearly half of the cases (43%). These results show the reemergence and changing of serotypes, the clinical association of respiratory adenovirus infections, and the molecular epidemiology of Ad7 genotypes in Taiwan during the past two decades. 相似文献
47.
目的探讨无精子症,严重少精子症和少、弱精子症患者的遗传缺陷与男性不育的关系。方法采用外周血染色体核型分析技术和Y染色体基因微缺失检测方法,对120例无精子症,严重少精子症和少弱精子的患者进行了遗传咨询。结果在被筛查患者中发现异常染色体核型13例,异常核型发生率为10.83%;而其Y染色体微缺失检测中存在AZFc/SPGY基因缺失31例,缺失率25.83%。结论染色体核型异常和Y染色体微缺失与精子生成障碍有直接逻辑关系。Y染色体AZFc/SPGY区域的微缺失是中国男性不育的重要原因,因此,中国男性不育症患者有必要进行Y染色体AZFc/SPGY微缺失的常规筛查。 相似文献
48.
Use of tuf sequences for genus-specific PCR detection and phylogenetic analysis of 28 streptococcal species 总被引:1,自引:0,他引:1 下载免费PDF全文
Picard FJ Ke D Boudreau DK Boissinot M Huletsky A Richard D Ouellette M Roy PH Bergeron MG 《Journal of clinical microbiology》2004,42(8):3686-3695
A 761-bp portion of the tuf gene (encoding the elongation factor Tu) from 28 clinically relevant streptococcal species was obtained by sequencing amplicons generated using broad-range PCR primers. These tuf sequences were used to select Streptococcus-specific PCR primers and to perform phylogenetic analysis. The specificity of the PCR assay was verified using 102 different bacterial species, including the 28 streptococcal species. Genomic DNA purified from all streptococcal species was efficiently detected, whereas there was no amplification with DNA from 72 of the 74 nonstreptococcal bacterial species tested. There was cross-amplification with DNAs from Enterococcus durans and Lactococcus lactis. However, the 15 to 31% nucleotide sequence divergence in the 761-bp tuf portion of these two species compared to any streptococcal tuf sequence provides ample sequence divergence to allow the development of internal probes specific to streptococci. The Streptococcus-specific assay was highly sensitive for all 28 streptococcal species tested (i.e., detection limit of 1 to 10 genome copies per PCR). The tuf sequence data was also used to perform extensive phylogenetic analysis, which was generally in agreement with phylogeny determined on the basis of 16S rRNA gene data. However, the tuf gene provided a better discrimination at the streptococcal species level that should be particularly useful for the identification of very closely related species. In conclusion, tuf appears more suitable than the 16S ribosomal RNA gene for the development of diagnostic assays for the detection and identification of streptococcal species because of its higher level of species-specific genetic divergence. 相似文献
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50.
The connexin26 gene ( GJB2) has been shown to be responsible for DFNB1 and DFNA3 (Autosomal Recessive Hereditary Nonsyndromic Deafness Locus 1 and Autosomal Dominant Hereditary Nonsyndromic Deafness Locus 3). Two hundred ten independently ascertained Chinese probands with nonsyndromic hearing loss (NSHL) were evaluated for mutations in GJB2, including 43 probands from families with more than one sib with NSHL, likely indicating dominant inheritance, and sporadic cases of NSHL, compatible with recessive inheritance. Of the 210 probands, 43 (20%) were homozygous or heterozygous for mutations in GJB2. Four different mutations were identified: 35delG, 109G-A, 235delC, and 299-300delAT. It was confirmed that GJB2 mutations are an important cause of hearing loss in this population. Of these four mutations, 235delC was the most prevalent at 93%; yet the 35delG mutation, which is the most common GJB2 mutation in Caucasian subjects (Europeans and Americans), was found in low frequency in the present study. It appears from our limited data and reports from other East Asians that 235delC is the most prevalent GJB2 mutation in these populations. GJB2 mutations are consistent with ethnic predilections. 相似文献