2-(2-Pyridyl)ethyl group: new type protecting group in the synthesis of DNA via phosphoramidite intermediates

2-(2-Pyridyl)ethyl group is a new type P-O protecting group for the synthesis of oligodeoxyribonucleotides by the phosphite triester method. This group is stable to alkali and acid conditions, and to be removed from internucleotidic bonds under mild conditions via two step procedures without any side reactions. Further we have found that bis(diisopropylamino)chlorophosphine is much more effective for the preparation of bis(diisopropylamino)alkoxyphosphines than various dichlorophosphines.     

Detection of IgG antibody to measles virus by radioimmunoassay technique

A highly sensitive procedure of solid-phase radioimmunoassay (RIA) was developed for the detection of measles IgG antibody. HeLa cells persistently infected with measles virus were used as a solid-phase antigen. This technique was applied to the detection of measles IgG antibody in patients with subacute sclerosing panencephalitis (SSPE) and multiple sclerosis. Normal subjects having experienced natural measles or measles vaccination and patients with various neurological diseases of non-virus nature were also examined as control groups. Measles antibody was detected at high titers in both the sera and cerebrospinal fluid of SSPE patients. Moreover, RIA/HI ratios of SSPE patients were significantly higher than those of normal subjects, suggesting the presence in the formers of antibodies to nucleocapsids at high titers as well as to viral envelopes. On the other hand, no significant difference was found in both RIA and HI titers between the sera of multiple sclerosis and those of various neurological diseases.     

Restriction endonuclease from Haemophilus gallinarum (HgaI) cleaves polyoma DNA at four locations.

A restriction endonuclease obtained from Haemophilus gallinarum (hgaI) cleaves polyoma DNA at four specific sites. Using the EcoRI, HindIII, and HpaII endonuclease restriction sites as reference, the four HgaI cleavage sites were mapped at 0.02, 0.14, 0.27, and 0.48 fractional lengths, clockwise, from the single EcoRI cleavage site.     

Superheated steam pyrolysis of biomass elemental components and Sugi (Japanese cedar) for fuels and chemicals

To develop a novel noncatalytic biomass refinery process that can be used as a portable process, superheated steam pyrolysis was investigated to produce both carbonized solid fuels and chemicals using a large-scale reactor. Individual biomass components and native biomass (Sugi, Japanese cedar) were pyrolyzed. Between 150 and 400 degrees C, the vaporizing fractions of cellulose, xylan, and kraft lignin were summarized using a numerical model. Cellulose was converted to glycolaldehyde, furfural, 5-hydroxymethyl furfural and levoglucosan, whereas xylan was converted to glycolaldehyde, furfural, and acetic acid. Kraft lignin produced a slight yield of phenol and guaiacol. The total vaporization fraction of Sugi and its vaporizing rate were explained sufficiently using a numerical model based on the weighted average of the vaporizing properties of the individual components. However, the yields of phenol, guaiacol, and acetic acid were underestimated, while the yields of furfurals and levoglucosan were overestimated. Possible synergetic effects among chemicals in the superheated steam pyrolysis of native biomass were also discussed.     

QTL-based analysis of leaf senescence in an indica/japonica hybrid in rice (Oryza sativa L.)

In order to identify quantitative trait loci (QTLs) for leaf senescence and related traits in rice (Oryza sativa L.), we developed two backcross populations, indica/japonica// japonica and indica/japonica//indica, using IR36 as the indica parent and Nekken-2 as the japonica parent. The QTLs were mapped using a set of simple sequence-repeat markers (SSRs) in the BC₁F₁ population. Senescence was characterized in these plants by measuring the leaf chlorophyll content 25 days after flowering (DAF), the reduction in chlorophyll content (the difference between the chlorophyll content at flowering and at 25 DAF), and the number of late-discoloring leaves per panicle at 25 DAF in five plants from each BC₁F₂ line. These plants were moved into a temperature-controlled growth cabinet at the time of flowering and allowed to mature under identical conditions. Eleven QTLs were detected in the two populations. The major of QTLs for senescence were found on the short arm of chromosome 6 and on the long arm of chromosome 9. Of these, one QTL on chromosome 6 and two on chromosome 9 were verified by confirming the effects of the genotypes on the phenotypes of the BC₁F₃ lines. The japonica parent was found to contribute to late senescence at all but one QTL. Based on a comparison of the effects of heterozygotes and homozygotes on the phenotypic values of each QTL genotype, we concluded that the differential senescence observed in the indica-japonica hybrid was not due to over-dominance; rather, it was the result of partial-dominance genes that were donated from either of the parents.     

Overexpression of the Escherichia coli catalase gene, katE, enhances tolerance to salinity stress in the transgenic indica rice cultivar, BR5

Salinity stress is a major limiting factor in cereal productivity. Many studies report improvements in salt tolerance using model plants, such as Arabidopsis thaliana or standard varieties of rice, e.g., the japonica rice cultivar Nipponbare. However, there are few reports on the enhancement of salt tolerance in local rice cultivars. In this work, we used the indica rice (Oryza sativa) cultivar BR5, which is a local cultivar in Bangladesh. To improve salt tolerance in BR5, we introduced the Escherichia coli catalase gene, katE. We integrated the katE gene into BR5 plants using an Agrobacterium tumefaciens-mediated method. The introduced katE gene was actively expressed in the transgenic BR5 rice plants, and catalase activity in T₁ and T₂ transgenic rice was approximately 150% higher than in nontransgenic plants. Under NaCl stress conditions, the transgenic rice plants exhibited high tolerance compared with nontransgenic rice plants. T₂ transgenic plants survived in a 200 mM NaCl solution for 2 weeks, whereas nontransgenic plants were scorched after 4 days soaking in the same NaCl solution. Our results indicate that the katE gene can confer salt tolerance to BR5 rice plants. Enhancement of salt tolerance in a local rice cultivar, such as BR5, will provide a powerful and useful tool for overcoming food shortage problems.     

Stimulation of arachidonic acid metabolism by a streptococcal preparation (OK-432) in rat peritoneal macrophages.

A streptococcal preparation OK-432 is reported to be an immunopotentiator and a potent antitumor agent. In order to elucidate the mechanism of biologic action, effects of OK-432 on arachidonic acid metabolism in rat peritoneal macrophages were investigated. Prostaglandin E2 production and release of radioactivity from [3H]arachidonic acid-labeled macrophages were found to be stimulated by OK-432 in a concentration-dependent manner (5 to 80 micrograms/ml). Heat-treatment of OK-432 further stimulated its effects. These stimulative effects on arachidonic acid metabolism by OK-432 were not observed in MDCK cells that have no phagocytotic activity. Furthermore, cytochalasin B treatment completely suppressed the stimulative effects induced by OK-432 in macrophages. These results strongly indicate that the stimulative effects by OK-432 on arachidonic acid metabolism are dependent on phagocytosis of OK-432 particles. Significance of stimulation of arachidonic acid metabolism in macrophages by OK-432 for its biological effects is discussed.     

Dopamine-dependent ectodomain shedding and release of epidermal growth factor in developing striatum: target-derived neurotrophic signaling (Part 2)

Epidermal growth factor (EGF) and structurally related peptides promote neuronal survival and the development of midbrain dopaminergic neurons; however, the regulation of their production has not been fully elucidated. In this study, we found that the treatment of striatal cells with dopamine agonists enhances EGF release both in vivo and in vitro. We prepared neuron-enriched and non-neuronal cell-enriched cultures from the striatum of rat embryos and challenged those with various neurotransmitters or dopamine receptor agonists. Dopamine and a dopamine D(1) -like receptor agonist (SKF38393) triggered EGF release from neuron-enriched cultures in a dose-dependent manner. A D(2) -like agonist (quinpirole) increased EGF release only from non-neuronal cell-enriched cultures. The EGF release from striatal neurons and non-neuronal cells was concomitant with ErbB1 phosphorylation and/or with the activation of a disintegrin and metalloproteinase and matrix metalloproteinase. The EGF release from neurons was attenuated by an a disintegrin and metalloproteinase/matrix metalloproteinase inhibitor, GM6001, and a calcium ion chelator, BAPTA/AM. Transfection of cultured striatal neurons with alkaline phosphatase-tagged EGF precursor cDNA confirmed that dopamine D(1) -like receptor stimulation promoted both ectodomain shedding of the precursor and EGF release. Therefore, the activation of striatal dopamine receptors induces shedding and release of EGF to provide a retrograde neurotrophic signal to midbrain dopaminergic neurons.     

Muscle mechanoreflex augments arterial baroreflex-mediated dynamic sympathetic response to carotid sinus pressure

Although the muscle mechanoreflex is one of the pressor reflexes during exercise, its interaction with dynamic characteristics of the arterial baroreflex remains to be quantitatively analyzed. In anesthetized, vagotomized, and aortic-denervated rabbits (n = 7), we randomly perturbed isolated carotid sinus pressure (CSP) using binary white noise while recording renal sympathetic nerve activity (SNA) and arterial pressure (AP). We estimated the transfer functions of the baroreflex neural arc (CSP to SNA) and peripheral arc (SNA to AP) under conditions of control and muscle stretch of the hindlimb (5 kg of tension). The muscle stretch increased the dynamic gain of the neural arc while maintaining the derivative characteristics [gain at 0.01 Hz: 1.0 +/- 0.2 vs. 1.4 +/- 0.6 arbitrary units (au)/mmHg, gain at 1 Hz: 1.7 +/- 0.6 vs. 2.7 +/- 1.4 au/mmHg; P < 0.05, control vs. stretch]. In contrast, muscle stretch did not affect the peripheral arc. In the time domain, muscle stretch augmented the steady-state response at 50 s (-1.1 +/- 0.3 vs. -1.7 +/- 0.7 au; P < 0.05, control vs. stretch) and negative peak response (-2.1 +/- 0.5 vs. -3.1 +/- 1.5 au; P < 0.05, control vs. stretch) in the SNA step response. A simulation experiment using the results indicated that the muscle mechanoreflex would accelerate the closed-loop AP regulation via the arterial baroreflex.     

The presence of the region on pBR322 that encodes resistance to tetracycline is responsible for high levels of plasmid DNA knotting in Escherichia coli DNA topoisomerase I deletion mutant.

Plasmid pBR322 DNA isolated from Escherichia coli DNA topoisomerase I deletion mutant DM800 is estimated to contain about 10% of the knotted forms (Shishido et al., 1987). These knotted DNA species were shown to have the same primary structure as usual, unknotted pBR322 DNA. Analysis of the knotting level of deletion, insertion and sequence-rearranged derivatives of pBR322 in DM800 showed that the presence of the region on pBR322 encoding resistance to tetracycline (tet) is required for high levels of plasmid knotting. When the entire tet region is present in a native orientation, the level of knotting is highest. Inactivating the tet promoter is manifested by a middle level of knotting. For deletion derivatives lacking various portions of the tet region, the level of knotting ranges from lowest to high depending on the site and length of the tet gene remaining. Inverting the orientation of tet region on the pBR322 genome results in a middle level of knotting. Deleting the ampicillin-resistance (bla)gene outside of its second promoter does not affect the level of knotting, if the entire tet gene remains. A possible mechanism of regulation of plasmid knotting is discussed.