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To clarify the cis-trans isomeric effect on the ultrafast DNA photo-cross-linking reaction of 3-cyanovinylcarbazole nucleoside ((CNV)K) in the DNA duplex, which gives a single photodimer on the reversed-phase HPLC chromatogram, the kinetics of the cis-trans photoisomerization of (CNV)K in double-stranded DNA was evaluated. Since the photoisomerization rate constant for cis to trans isomerization in double-stranded DNA was significantly larger than that for trans to cis isomerization, and the thermodynamic stability of the trans isomer was higher than that of the cis isomer, it was strongly suggested that the trans isomer of (CNV)K is a reactive species of the photo-cross-linking reaction. (1)H-(1)H NOESY analysis of the photoadduct consisting of (CNV)K and T also supported the trans-mediated photo-cross-linking reaction of (CNV)K. By using this ultrafast photo-cross-linking reaction for the molecular beacon-based SNPs typing, four individual Japanese rice strains were clearly distinguishable with simple photoirradiation and fluorescence imaging using double-stranded target DNAs.

Objective: This was an open-labeled, prospective, control study to determine the efficacy of methotrexate (MTX) for improving serological abnormalities in patients with systemic lupus erythematosus (SLE)., Methods: Thirty patients with a low serum complement and/or high anti-double-stranded DNA (dsDNA) antibody levels during a prednisolone taper received MTX orally at a dose of 7.5 mg/week over 12-18 months (MTX group). Eighteen SLE patients were selected as controls (control group). At the time of entrance into the study, all patients were receiving <15 mg/day of prednisolone. The C3, C4, and immunoglobulin (Ig)G, IgA, and IgM levels were measured using a turbidimetric immunoassay. The anti-dsDNA antibody level was measured using the Farr assay., Results: Significant increases in C3 and C4 levels and significant decreases in anti-dsDNA antibody, IgG, IgA, and IgM levels from baseline were observed for 3-18 months after the trial in the MTX group but not in the control group. At the end of the study, C3 and/or C4 levels in 96.7% of the MTX patients and 33.3% of the control patients were normalized or elevated (p = 0.0001), and anti-dsDNA antibody levels were normalized or lowered in 24 of the 26 MTX patients (92.3%) and in 50.0% of the control patients (p = 0.0022). In addition, a significant reduction in SLE Disease Activity Index (SLEDAI) score and a prednisolone-sparing effect were observed for the MTX group but not the control group. A significant increase in mean corpuscular volume of red blood cells, which is indicative of an anti-folic-acid metabolic disorder induced by MTX, was observed only for patients in the MTX group. Five patients (16.7%) discontinued MTX treatment because of disease flare, and another three (10.0%) discontinued due to treatment side effects., Conclusion: MTX appears to be effective for improving serological abnormalities frequently observed before disease flares in SLE patients on a prednisolone taper.

To develop a covalent and specific labeling method for single- and double-stranded plasmid DNA, photoresponsive oligonucleotide containing 3-cyanovinylcarbazole nucleoside was adopted. Single- and double-stranded plasmid DNA was successfully labeled/de-labeled with Cy3 and/or biotin by photoirradiation.

We describe a photoreversible DNA end capping via 3-cyanovinylcarbazole nucleoside. Doubly end-capped oligodeoxynucleotide (ODN) exhibits increased stability against snake venom phosphodiesterase and shows high thermal stability.

We present a new type of nanoparticle-based DNA sensor using surface-enhanced Raman scattering (SERS) on gold nanoparticle (Au NP) aggregates formed by DNA photoligation. The DNA sensor exploits the photoligation reaction between oligodeoxynucleotides (ODNs) attached to the surfaces of Au NPs in the presence of target DNA (T-DNA). When hybridization takes place between the ODNs and T-DNA, Au NPs are covalently crosslinked to form aggregates via photoligation. Once the NP aggregates are formed, the interspace between Au NPs in the aggregate act as a stable "hot spot", and a SERS signal from the Raman-active molecules (sodium cacodylate) present in the hot spot is easily and sensitively detected. In contrast, a SERS signal is not detected if the hybridization is unsuccessful, because the stable hot spot does not form. This DNA sensor does not require an enzymatic reaction, fluorescent dye, precise temperature control, or complicated operating procedures.

We describe a novel ultrafast reversible DNA interstrand photo-cross-linking reaction via 3-cyanovinylcarbazole nucleoside ( (CNV)K). Oligodeoxynucleotide (ODN) containing (CNV)K can be photo-cross-linked by irradiation at 366 nm for 1 s, and the photo-cross-linked ODN can be split by irradiation at 312 nm for 60 s.

We describe a light-controlled template-directed reversible DNA photoligation via carbazole-tethered 5-carboxyvinyluracil. Carbazole-tethered 5-carboxyvinyl-2'-deoxyuridine-containing oligodeoxynucleotide (ODN) can be ligated by irradiation at 366 nm in the presence of template ODN, and the ligated ODN can be split by irradiation at 366 nm in the absence of template ODN.

We describe a novel reversible DNA interstrand photocrosslinking reaction via the artificial nucleoside (X). Oligodeoxynucleotide (ODN) containing X can be photocrosslinked by irradiation at 366 nm for 1 s, and the photocrosslinked ODN can be split by irradiation at 312 nm for 60 s.

We describe a simple and inexpensive SNP typing method by using sequence specific interstrand photocrosslinking via p-carbamoylvinyl phenol nucleosides. Interstrand photocrosslinking showed a high degree of single nucleotide specificity as high as 10(3)-fold and more, and can be used in the diagnostic detection of DNA sequences.

We describe a highly efficient method for reversible photocircularization of oligonucleotide (ODN) on a double-stranded DNA template: 5-carboxyvinyl-2'-deoxyuridine-containing ODN was reversibly circularized around the target sequence of the double-stranded plasmid DNA resulting in formation of a catenated plasmid.

We report the nonenzymatic terminal labeling of oligoribonucleotide (ORN) by using template-directed photoligation through 5-carboxyvinyl-2'-deoxyuridine ((CV)U) with high selectivity.

We describe a highly efficient template-directed photoligation of oligodeoxynucleotides (ODNs) through 5-carbamoylvinyl-2'-deoxycytidine ((CV)C). When an ODN containing (CV)C at the 5' end was photoirradiated with an ODN containing a pyrimidine base at the 3' end in the presence of template DNA, efficient photoligation was observed without any byproduct formation. Single nucleotide differences can be successfully distinguished by using photoligation-based DNA chip assay. [structure: see text]

We report that deamination coupled with 5-carboxyvinyldeoxyuridine-mediated photobranching causes the heat-induced transition of cytosine to uracil with high efficiency without any side reaction.

The herpes simplex virus (HSV) amplicon vector is a versatile plasmid-based gene delivery vehicle with a large transgene capacity (up to 150 kb) and the ability to infect a broad range of cell types. The vector system was originally developed by Frenkel and her colleagues in 1980. Ever since, a great deal of effort by various investigators has been directed at minimizing the toxicity associated with the inevitable contamination by helper virus. In 1996, Fraefel and his colleagues successfully devised a cosmid-based packaging system that was free of contamination by helper virus (so-called helper virus-free packaging), which utilized as helper a set of 5 overlapping cosmid clones that covered the entire HSV genome, which lacked the DNA packaging/cleavage signals. With the helper virus-free system, broader applications of the vector became possible. Cloning of the entire HSV genome in bacteria artificial chromosome (BAC) plasmids enabled stable maintenance and propagation of the helper HSV genome in bacteria. It also allowed for the development of BAC-based helper virus-free packaging systems. In this article, we review various versions of DNA-based methods to prepare HSV amplicon vectors free of helper virus contamination. We also examine recent advances in vector design, including methods of vector construction, hybrid amplicon vectors, and the infectious BAC system. Future directions in improving packaging systems and vector designs are discussed.

We report the photoinduced repair of a thymine dimer incorporated in a DNA duplex via oligodeoxynucleotide (ODN) containing carbazole nucleoside (K). The occurrence of an electron transfer between K and thymine dimer is evidenced by fluorescence quenching measurements. K acts as a good electron donor for the photoinduced repair of a thymine dimer.

We previously reported an efficient and reversible template-directed photoligation using 5-carboxyvinyl-2'-deoxyuridine ((CV)U)-containing ODN at the 5'-terminal. This method forms d(T-(CV)U) as a cyclobutane pyrimidine dimer (CPD) analogue between 3'-terminal thymidine and 5'-terminal (CV)U of two oligodeoxynucleotides (ODNs). In this study, we performed PCR using a DNA template containing d(T-(CV)U). And then, we disclosed that two adenines were incorporated opposite the d(T-(CV)U).

[reaction: see text] We describe an efficient template-directed photoligation of oligodeoxynucleotides (ODNs) using alpha-5-cyanovinyldeoxyuridine (alpha(C)U). An efficient photoligation was produced by photoirradiation of an ODN containing alpha(C)U at the 3' end with an ODN containing thymine at the 5' end in the presence of a template ODN. This photoligation method is a new and efficient way to synthesize branched ODNs.

A modified oligodeoxynucleotide (ODN) containing N(3)-methyl-5-cyanovinyl-2'-deoxyuridine reacts by photoirradiation at 366 nm with an adenine residue of a complementary template ODN to yield an end-capped ODN in 87% yield.

We report a novel interstrand photocrosslinking of oligodeoxynucleotides (ODNs). In this system, a modified ODN containing p-carbamoylvinyl phenol nucleoside reacts by photoirradiation at 366 nm with adenine residue of a complementary template ODN to yield a crosslinked ODN in 97% yield.

We report a novel interstrand photocrosslink of oligodeoxynucleotides (ODNs). In this system, a modified ODN containing 3-carbonylvinyl phenol nucleoside reacts by photoirradiation at 366 nm with adenine residue of a complementary template ODN to yield a crosslinked ODN.

An efficient and reversible Template-directed Photoligation of oligodeoxynucleotides (ODNs) using alpha-5-cyanovinyldeoxyuridine (alpha-(CNV)U) was described. When ODN containing alpha-(CNV)U at the 3' end was photoirradiated with ODN containing thymidine at the 5' end in the presence of a template ODN, an efficient photoligation was observed. The ligated ODN can be split site-selectively to regenerate parent ODN by photoirradition at the 312 nm.

The photochemical DNA computing of 5-vinyluracil ((V)U) containing oligodeoxynucleotides (ODNs) tethered multiple "DNA words" is demonstrated. A new MARK and UNMARK operation based on the (V)U mediated reversible DNA photoligation has been developed for multiple-word DNA computing. The utility of this operation for DNA computing is demonstrated by solving a small Satisfiability problem (SAT problem) in which information was encoded in three tandem words.

N:-Nitrosopyrrolidine (NPYR) is carcinogenic in rodents and undergoes alpha-hydroxylation upon microsomal CYP450 metabolism, giving rise to mutations. Previously, we reported the direct mutagenicity of NPYR, under ultraviolet A (UVA) irradiation, towards Salmonella typhimurium and phage M13mp2. In the present study, we measured the formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) in a replicative form of M13mp2 DNA exposed to NPYR plus UVA. Formation of 5-hydroxy-2'-deoxycytidine in calf thymus DNA treated with NPYR plus UVA was also observed. Singlet oxygen is likely to account for the formation of 8-oxodGuo. We analyzed the spectrum of mutations in lacZalpha of M13mp2 phages produced on transfecting Escherichia coli with the replicative form of phage DNA that had been treated with NPYR plus UVA. The role of oxidative DNA damage in mutagenesis was explored using mutM-proficient and -deficient E.coli strains as the hosts. A higher level of mutation was observed with the mutM-deficient host than with the -proficient host. Base substitutions at GC pairs predominated in both mutM-proficient and -deficient hosts. With the mutM-deficient host, we observed an overall increase in the percentage of GC-->TA transversions. In addition we noted that there were fewer GC-->AT transitions than in the mutM-proficient host. With these hosts, different hot spots were observed and a new GC-->TA hot spot was produced. The formation of 8-oxodGuo in DNA, which is known to induce GC-->TA transversion, may contribute to mutagenesis by NPYR plus UVA.