Development of a simple and efficient method for transformation of buckwheat plants (Fagopyrum esculentum) using Agrobacterium tumefaciens

Apical meristems of seedlings of buckwheat (Fagopyrum esculentum var. Shinano No. 1) were pricked with a needle and inoculated with Agrobacterium tumefaciens (LBA4404, pBI121). The inoculated seedlings were grown to maturation and allowed to pollinate randomly to set the seeds (T1 plants). The transformation efficiency of the T1 plants was estimated by germination in the presence of geneticin (20 microg/ml) and by detection of beta-glucuronidase (GUS) gene with PCR, indicating that 36% and 70% of the T1 plants were transformed, respectively. Four plants taking on a mutated morphology were selected from T1 plants which were transformed with the method using A. tumefaciens harboring a modified pBI121 for plasmid rescue. Southern blot analysis of plasmids rescued from the 4 T1 plants demonstrated that each plasmid contained a different flanking DNA of the buckwheat genome, an evidence that T-DNA was integrated in different sites of the genomic DNA among the 4 T1 plants.     

Nucleotide sequence and copy control function of the extension of the incI region (incI-b) of Rts 1

An Rts1 derivative, pTW20, contains three incompatibility (inc) regions, incI-a (incI in previous studies), incII, and newly determined incI-b loci. By restriction analysis, we have located the incI-b adjacent to the incI-a region on the pTW20 map. Nucleotide sequence analysis of the minimal incI-b region revealed the presence of four repeated sequences, each consisting of 18 bp, which is similar to the incI-a and incII repeats existing on mini-Rts1. All four repeating units were required for expression of a strong incompatibility. In addition, RepA protein, essential for the replication of Rts1, bound specifically to the repeated sequences, suggesting that the repeats would titrate out RepA protein as do incI-a and IncII. Insertion of the incI-b to a mini-Rts1 plasmid in a natural arrangement decreases the copy number of mini-Rts1 to the same level as that of mini-F. The incI-a and incI-b might be a single constituent in incompatibility and copy number control of Rts1.     

Metaphase chromosomes of four species of the Drosophila nasuta subgroup

Metaphase chromosomes of four species of the Drosophila nasuta subgroup, D. nasuta, D. kepulauana, D. kohkoa, and D. neveifrons, were analysed by the C banding method to clarify the chromosomal differentiation during speciation. Four species were different in heterochromatic regions of their chromosomes, especially microchromosomes of the fourth chromosomes and the Y chromosomes. Intraspecific variations of heterochromatic regions were also found among isofemale lines from various localities of D. nasuta, D. kepulauana, and D. kohkoa. Intraspecific variations of heterochromatic regions were not different from interspecific variations morphologically. From these results, the evolutionary process of heterochromatic regions was discussed.     

Expression of a vacuolar protein (VP24) in anthocyanin-producing cells of sweet potato in suspension culture.

VP24, an abundant protein of 24 kD, was found to accumulate in the anthocyanin-containing vacuoles of cells of sweet potato (Ipomoea batatas) in suspension culture. Light-induced expression of VP24 was analyzed by immunoblotting in three different cell lines that produced anthocyanins at different rates. The expression of VP24 was closely correlated with the accumulation of anthocyanin in these cell lines. Immunocytochemical detection of VP24 with specific antibodies on thin sections showed that VP24 was localized in the intravacuolar pigmented globules (cyanoplasts) in the anthocyanin-containing vacuoles and not in the tonoplast. No VP24 immunogold labeling was detected in the vacuoles of the cell line that does not produce anthocyanin. We suggest that VP24 may be involved in the formation of the cyanoplast via an interaction with anthocyanin, and that it may play an important role in the trapping in vacuoles of large amounts of anthocyanins that have been transported into these vacuoles.     

Context Dependent Effect of Landscape on the Occurrence of an Apex Predator across Different Climate Regions

In studies of habitat suitability at landscape scales, transferability of species-landscape associations among sites are likely to be critical because it is often impractical to collect datasets across various regions. However, limiting factors, such as prey availability, are not likely to be constant across scales because of the differences in species pools. This is particularly true for top predators that are often the target for conservation concern. Here we focus on gray-faced buzzards, apex predators of farmland-dominated landscapes in East Asia. We investigated context dependency of “buzzard-landscape relationship”, using nest location datasets from five sites, each differing in landscape composition. Based on the similarities of prey items and landscape compositions across the sites, we determined several alternative ways of grouping the sites, and then examined whether buzzard-landscape relationship change among groups, which was conducted separately for each way of grouping. As a result, the model of study-sites grouping based on similarities in prey items showed the smallest ΔAICc. Because the terms of interaction between group IDs and areas of broad-leaved forests and grasslands were selected, buzzard-landscape relationship showed a context dependency, i.e., these two landscape elements strengthen the relationship in southern region. The difference in prey fauna, which is associated with the difference in climate, might generate regional differences in the buzzard-landscape associations.     

Qualitative and quantitative analysis of cytologic assessment of astrocytoma, oligodendroglioma and oligoastrocytoma

OBJECTIVE: To evaluate the usefulness of intraoperative cytology for differential diagnoses of astrocytoma, oligodendroglioma and oligoastrocytoma. STUDY DESIGN: Qualitative analysis of cytologic features of the 3 brain tumors was conducted using intraoperative touch or squash preparations that were stained with the Papanicolaou method, targeting the cellular density, cytoplasmic and nuclear profiles and blood vessel morphology. In addition, we attempted a computer-assisted image analysis of tumor cell nuclei and compared the results with qualitative observations. RESULTS: Astrocytomas were characterized by many fibrillary cytoplasmic processes and large, irregular nuclei. Oligodendrogliomas were characterized by small, round nuclei and a fine, delicate capillary network. In both tumors of a higher grade, anaplastic large nuclei and proliferating endothelial cells were noted. Oligoastrocytomas showed combined cytologic profiles of astrocytomas and oligodendrogliomas. Quantitative studies suggested that nuclei of oligodendroglial tumors were significantly rounder than those of astrocytomas. In general, the quantitative results were consistent with the qualitative observations. CONCLUSION; Cytologic evaluation using touch or squash preparations is of great help for intraoperative differential diagnosis of astrocytoma, oligodendroglioma and oligoastrocytoma. Cytologic as well as histologic assessment should be conducted at the intraoperative diagnosis of these tumors.     

Chemotherapy for enterohemorrhagic Escherichia coli O157:H infection in a mouse model

The aim of this study is to evaluate the therapeutic effect of the antimicrobial agents, fosfomycin (FOM), minocycline (MINO), kanamycin (KM) and norfloxacin (NFLX) in the enterohemorrhagic Escherichia coli (EHEC) infected mouse model which we established previously (Infect. Immun. 62 (1994) 3447-3453). Each of the antimicrobial agents, 1/16 LD(50), was given to the mice per os (p.o. ) or intraperitoneally (i.p.) for 3 days after bacterial inoculation and then we observed their mortality rate for 2 weeks. The mortality rates of mice administered with MINO (p.o./i.p.), KM (p.o.), NFLX (p. o./i.p.) were significantly lower than those of the control group. Both the bacterial number and VT2c level in the feces of the FOM group were lower than those of the NFLX group on day 1, but reversed on day 3. In an in vitro experiment, each of the four drugs in combination with mitomycin C (MMC) caused a more significant decrease in the bacterial number than sole MMC, and they consequently indicated the suppressive effect on the release of VT2c.