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1.
Site-directed cleavage of RNA.   总被引:18,自引:12,他引:6       下载免费PDF全文
Using complementary chimeric oligonucleotides containing deoxyribonucleotides and 2'-O-methylribonucleotides (1), enzymatically synthesized RNA (90 mer) were cleaved at a single site with Escherichia coli RNaseH, either at a hairpin loop or at a stem region. Especially, site-specific cleavage occurred in even the target region being enclosed within a stable, base-paired stem. The method is proved to be generally applicable to RNA containing secondary structures.  相似文献
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NMR studies of a DNA containing 8-hydroxydeoxyguanosine.   总被引:13,自引:6,他引:7       下载免费PDF全文
The effects of hydroxylation at the C8 of a deoxyguanosine residue in DNA were studied by NMR analysis of a self-complementary dodecanucleotide, d(C1-G2-C3-oh8G4-A5-A6-T7-T8-C9-G10-C11-G12), which has an 8-hydroxy-2'-deoxyguanosine (oh8dG) residue at the 4th position. NMR data indicate that the 8-hydroxyguanine (oh8G) base takes a 6,8-diketo tautomeric form and is base-paired to C with Watson-Crick type hydrogen bonds in a B-form structure. The thermal stability of the duplex is reduced, but the overall structure is much the same as that of the unmodified d(CGCGAATTCGCG) duplex. The structural changes caused by 8-hydroxylation of the deoxyguanosine, if any, are localized near the modification site.  相似文献
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Y Hayase  H Inoue  E Ohtsuka 《Biochemistry》1990,29(37):8793-8797
In order to cleave RNA at specific positions in Escherichia coli formylmethionine tRNA, RNase H and complementary chimeric oligonucleotides consisting of DNA and 2'-O-methyl-RNA (Inoue et al. (1987) FEBS Lett. 215, 327] were used. Specific cleavages in the D loop, anticodon loop, T psi C loop, anticodon stem, and acceptor stem were investigated. Virtually unique hydrolyses with RNase H were observed at the T psi C loop, anticodon stem, and acceptor stem when relatively longer chimeric oligonucleotides (20-mer) were used. An efficient cleavage at the anticodon was obtained with a chimeric 13-mer when the higher structure of the tRNA was broken by hybridization with a 20-mer at the acceptor as well as the T psi C stem region. It was found that stabilities of hybrids with chimeric oligonucleotides and the presence of minor nucleosides affect the cleavage of tRNA by this approach.  相似文献
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Six nonaribonucleotides containing the 5'-splice site, one complementary nonamer and an octadecamer containing the 3'-splice site have been synthesized on a polymer support using the phosphoro-p-anisidate method. A 5'-linked 2'-O-tetrahydrofuranyl-N-protected nucleoside 3'-(o-chlorophenyl)phosphoro-p-anisidate was used as the starting nucleotide, and the chain elongated in the 3'-direction by removing the p-anisidate protecting group with isoamyl nitrite under neutral conditions. The octadecamer has been synthesized using dinucleotide blocks and a 3'-terminal trinucleotide.  相似文献
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The RHO1 gene encodes a yeast homolog of the mammalian RhoA protein. Rho1p is localized to the growth sites and is required for bud formation. We have recently shown that Bni1p is one of the potential downstream target molecules of Rho1p. The BNI1 gene is implicated in cytokinesis and the establishment of cell polarity in Saccharomyces cerevisiae but is not essential for cell viability. In this study, we screened for mutations that were synthetically lethal in combination with a bni1 mutation and isolated two genes. They were the previously identified PAC1 and NIP100 genes, both of which are implicated in nuclear migration in S. cerevisiae. Pac1p is a homolog of human LIS1, which is required for brain development, whereas Nip100p is a homolog of rat p150(Glued), a component of the dynein-activated dynactin complex. Disruption of BNI1 in either the pac1 or nip100 mutant resulted in an enhanced defect in nuclear migration, leading to the formation of binucleate mother cells. The arp1 bni1 mutant showed a synthetic lethal phenotype while the cin8 bni1 mutant did not, suggesting that Bni1p functions in a kinesin pathway but not in the dynein pathway. Cells of the pac1 bni1 and nip100 bni1 mutants exhibited a random distribution of cortical actin patches. Cells of the pac1 act1-4 mutant showed temperature-sensitive growth and a nuclear migration defect. These results indicate that Bni1p regulates microtubule-dependent nuclear migration through the actin cytoskeleton. Bni1p lacking the Rho-binding region did not suppress the pac1 bni1 growth defect, suggesting a requirement for the Rho1p-Bni1p interaction in microtubule function.  相似文献
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A multifunctional calmodulin-dependent protein kinase in the canine cardiac cytosol was purified to near homogeneity. The purified enzyme inactivated glycogen synthase by means of phosphorylation. The enzyme also phosphorylated phospholamban and several other proteins. In view of its physicochemical properties and substrate specificity, the enzyme differed from myosin light chain kinase and phosphorylase kinase, and was considered to belong to a class of similar calmodulin-dependent protein kinases from brain, liver, and skeletal muscle. The results suggest that the enzyme mediates multiple Ca2+-dependent functions in the heart.  相似文献
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The biological and serological properties of two Japanese barley mild mosaic virus (BaMMV) strains (BaMMV-Kal and BaMMV-Nal) and a German BaMMV strain (BaMMV-M) were compared. Mechanical inoculation experiments showed that these three strains differed from one another in their ability to infect specific barley cultivars. BaMMV-Kal and BaMMV-M caused similar symptoms, but BaMMV-Nal clearly differed from them in its symptoms on some barley cultivars. The three BaMMV strains efficiently infected barley plants at 15°C, whereas at 20°C BaMMV-Kal and BaMMV-M also infected many plants but BaMMV-Nal infected only a few. BaMMV-Kal and BaMMV-M were indistinguishable by ELISA, while BaMMV-Nal was distinguished from both. The biological and serological variability reported shows that BaMMV occurs as two distinct strains in Japan.  相似文献
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