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51.
We used the polymerase chain reaction (PCR) and direct DNA sequencing to study genetic variation within and among populations of Atlantic cod, Gadus morhua , in the western North Atlantic. In a 307 bp region of the mitochondrial cytochrome b gene, 24 variable nucleotide positions define 24 genotypes, which differ by from one to six nucleotide substitutions. Greenland cod ( G. ogac ) differs from the most similar G. morhua genotype by an additional 12 nucleotide substitutions. Silent transitions dominate both intra- and interspecific comparisons, however four nucleotide substitutions within morhua result in amino acid replacements. Direct sequencing of DNA reveals substantially more of the genetic variation that exists within and between species than do previous indirect methods based on restriction fragment length polymorphisms, and thus has far greater potential to quantify such differences as may exist among fish stocks. Preliminary experiments also indicate that automation of DNA sequencing provides an efficient, rapid, and accurate means for detection of genetic variation in natural populations offish. 相似文献
52.
53.
Jean- -François Laliberté Olivier Nicolas Serge Durand Rolf Morosoli 《Plant molecular biology》1992,18(3):447-451
The xylanase gene from Cryptococcus albidus contains seven introns. Genomic and cDNA clones under the control of the CaMV 35S promoter were transferred into tobacco plants using Agrobacterium-mediated cell transformation. The genes were transcribed and the mRNAs were amplified by the polymerase chain reaction using primers on each side of the intron region. About 90% of the amplification products from plants transformed with the genomic clone corresponded to the size of the pre-mRNA (1.2 kb) and 10% represented the spliced product (0.85 kb). The 0.85 kb fragment was cloned and sequenced and the result indicated that the introns from the xylanase gene were accurately spliced by the plant cells. 相似文献
54.
PCR analysis of dystrophin gene mutation and expression. 总被引:2,自引:0,他引:2
J S Chamberlain N J Farwell J R Chamberlain G A Cox C T Caskey 《Journal of cellular biochemistry》1991,46(3):255-259
55.
An assay procedure was developed that allowed the first reproducible measurement of DNA polymerase activity in all developmental stages of Drosophila melanogaster. Evidence is presented that the same enzymatic species is present in extracts of embryos, pupae, and adults of both sexes and that this activity has many properties similar to vertebrate α-polymerases. Polymerase activity per individual is low in embryos and rises steadily through larval instars, reaches a peak in early pupae, declines through the late pupal period, and remains low in newly eclosed adults of both sexes. A dramatic increase is observed in adult females as mature oocytes are formed. This pattern of enzyme activity is completely coincident with changes in DNA levels during development, and suggests that the Drosophila enzyme, like vertebrate α-polymerases, functions in cellular DNA replication. Two mutagen-sensitive mutants, deficient in both replication on undamaged templates and postreplication repair, were found to have normal levels of this α-polymerase activity. Our results suggest that a single enzymatic species of α-polymerase holoenzyme exists in Drosophila and is common to all developmental stages of this organism. 相似文献
56.
Summary Tritiated -amanitin has been used as a specific and sensitive probe to estimate the number of RNA polymerase B molecules in isolated nuclei, chromatin and nucleoids, obtained from macroplasmodia ofPhysarum polycephalum. During mitosis (metaphase±10 min) there is at least 10-fold less RNA polymerase B than at all phases of the cell cycle, even if DNA replication has been blocked in vivo. It is concluded that many of the RNA polymerase B molecules leave the chromatin during decondensation of the chromosomes in telophase of the synchronous nuclear division ofPhysarum. 相似文献
57.
58.
D. L. Fine L. O. Arthur L. J. T. Young 《In vitro cellular & developmental biology. Plant》1976,12(10):693-701
Summary Several cell culture factors were found to influence in vitro expression of mouse mammary tumor virus (MMTV) in the mouse
adenocarcinoma cell line Mm5mt/c1. Cells were propagated in a variety of commercially available cell culture media to which dexamethasone (DXM) was added as
a stimulator of MMTV production. Culture seeding density, culture medium type, and glucose concentration each influenced MMTV
production when expressed on a per cell basis. Maximum cell growth occurred in cultures grown in RPMI-1640 medium containing
insulin. Those media which provided good cell growth were not necessarily optimal for virus expression. Addition of insulin
did not stimulate MMTV synthesis although dexamethasone alone was stimulatory in all media used; however, maximum MMTV expression
was obtained when dexamethasone and insulin were used in concert. Equivalent levels of MMTV-specific cell membrane antigen,
MMTV-specific protein, and virus particles were produced at incubation temperatures of 32°, 34° or 37° C; however, higher
levels of virus-related RNA-dependent DNA polymerase (RDDP) activity were recovered from cultures incubated at 32° and 34°
C than at 37° C. Decreased levels of RDDP were attributed to enzyme thermolability at 37° C incubation.
Research sponsored by the National Cancer Institute under Contract No. N01-CO-25423 with Litton Bionetics, Inc., and Contract
No. N01-CP-33253 with the University of California. 相似文献
59.
60.
丙型肝炎病毒RNA打点杂交检测方法同RT-PCR方法的比较 总被引:1,自引:0,他引:1
采用HCV基因组结构区C区cDNA探针和非结构区NS3-4区cDNA探针,建立了用打点杂交(dotblothybridization)检测血清中HCVRNA的方法,同采用HCV基因组5’端非编码区的一对寡核苷酸引物通过逆转录-聚合酶链式反应(RT-PCR)检测血清中HCVRNA的方法相比较,发现两种方法都能快速早期和特异地检出血清中HCVRNA,但RT-PCR法敏感性优于RNA打点杂交法。对于无血清学指标的慢性NANB肝炎病人的诊断,可采用这两种方法。这两种方法的敏感性在很大程度上依赖于引物和探针的敏感性,以及RNA提取方法。RT-PCR法适用于诊断病毒血症和复制,打点杂交法适用于研究HCVRNA量的变化,对治疗的评价,以及为实验筛选较高滴度的HCVRNA阳性样本。 相似文献