Nucleic acids symposium series最新文献

The 1'alpha-phenylselenouridine derivative (4) was successfully synthesized via enolization at the 1'-position of the 3',5'-O-TIPDS-2'-ketouridine (1). After the introduction of a vinylsilyl tether as an intramolecular radical acceptor at the 2'-hydroxy group of 4, its atom-transfer radical cyclization reaction, followed by the treatment with TBAF gave 1'alpha-vinyluridine derivative (10). Using this procedure, 1'alpha-vinyluridine (11) and -cytidine (14) were successfully synthesized.

Synthetic studies on phosphodiester, phosphorothioate, and phosphorodithioate-linked oligonucleotides in terms of 2-(levulinyloxymethyl)-5-nitrobenzoyl (LMNBz) group as the base-labile protecting group for the 5'-hydroxyl groups of nucleoside 3'-H-phosphonate and -H-phosphonothioate derivatives, are described.

Properties of 2'-O-methyloligoribonucleotides containing two consecutive 2'-O-(1-pyrenylmethyl)uridine were investigated as a fluorescent probe to search the single strand regions of RNA. The bis-pyrene-labeled 2'-O-methyloligoribonucleotide (OMUpy2) induced the formation of pyrene dimer upon hybridization with the complementary oligoribonucleotides and showed remarkable appearance of broad structureless fluorescence at 480 nm. Contrarily, when OMUpy2 was hybridized with the complementary oligodeoxyribonucleotides, such enhancement of fluorescence was scarcely observed. When various OMUpy2 were applied to E. coli 5S-rRNA, the fluorescence intensity at 480 nm was varied in a sequence specific manner.

In our previous studies, it was demonstrated that the activity of a ribozyme in vivo was governed by several parameters, which include a high level-expression of ribozyme, the intracellular stability of the ribozyme and colocalization of the ribozyme with its target RNA in the same cellular compartment. To generate ribozymes with significant activity in vivo, we have developed a ribozyme-expression system based on a human tRNA(Val) promoter. Our tRNA-embedded ribozymes produced by our ribozyme-expression system remain relatively stable in cultured cells with half-lives longer than 30 min. Moreover, tRNA-ribozymes with a cloverleaf structure were efficiently exported from the nucleus to the cytoplasm, where they would effectively cleave target RNAs. In the present study, we investigated the relationship between the secondary structure of the tRNA-ribozymes and the transport efficacy of them in mammalian cells by using a screening system in vivo. Furthermore, we also investigated the mechanism of the export of tRNA-embedded ribozymes both in mammalian cells and in Xenopus oocytes.

It is found that T4 phage DNA complexed with histone H1 assembled into a string-of-bead structure, when the complex is prepared by a gentle diluting procedure from a high salt solution (2 M NaCl) to a low salt solution (50 mM NaCl). We used fluorescence microscopy to perform the real-time observation on formation and motion of a string-of-bead structure. Spatial histone H1 distribution on the DNA-H1 complex is observed by immuno-fluorescence microscopy.

A novel probe (Smart probe) has been developed for nucleic acid detection. The smart probe is an oligodeoxyribonucleotide carrying a fluorophore and an intercalator internally. Fluorescence of the smart probe is quenched by the intercalator in the absence of target sequence. While upon hybridization the probe emits greater fluorescence due to the interference of quenching by intercalation. The smart probe has been shown to recognize a single base mismatch in the double-stranded form without utilizing thermal stability difference of hybrids.

Genome of A. tumefaciens contains a linear and a circular chromosome. As an initial step of elucidating the structural and functional genomics of this bacterium, linkage map of the left region of its linear chromosome was constructed. Total genomic libraries of A. tumefaciens MAFF301001 were constructed in BAC vectors namely, pFOS1 and pBeloBAC11. Upon construction of sub-libraries, minimum overlapping clones needed to cover the left region was determined. So far, four contigs have been assembled with a total of 19 overlapping clones. Detailed EcoRI physical map of contig III was constructed and it covers a 110 kb region of the Pme5 fragment of the linear chromosome. Seven end regions of the linking clones were partially sequenced but no gene existence was determined due to low homology.

In the RNA of hyperthermophiles, which grow optimally between 80 degrees C and 106 degrees C, posttranscriptional modification has been identified as a leading mechanism of structural stabilization. Particularly in the Archaeal evolutionary domain these modifications are expressed as a structurally diverse array of modification motifs, many of which include ribose methylation. Using mass spectrometric techniques we have examined the posttranscriptional modifications in unfractionated tRNA from the remarkable organism Pyrolobus fumarii, which grows optimally at 106 degrees C, but up to 113 degrees C (Blöchl et al. (1997), Extremophiles, 1, 14-21). Twenty-six modified nucleosides were detected, 11 of which are methylated in ribose. A new RNA nucleoside, 1,2'-O-dimethylguanosine (m1Gm) was characterized and the structure confirmed by chemical synthesis.

Modified DNA carrying an azobenzene was successfully applied to the photo-regulation of DNA/RNA hybridization. When the azobenzene was isomerized from trans- to cis-form on UV-irradiation, the melting temperature of the duplex was significantly lowered. This process was totally reversible so that the Tm increased by cis-->trans isomerization induced by visible light irradiation.