Recent genetic analysis showed that phototropins (phot1 and phot2) function as blue light receptors in stomatal opening of Arabidopsis thaliana, but no biochemical evidence was provided for this. We prepared a large quantity of guard cell protoplasts from Arabidopsis. The immunological method indicated that phot1 was present in guard cell protoplasts from the wild-type plant and the phot2 mutant, that phot2 was present in those from the wild-type plant and the phot1 mutant, and that neither phot1 nor phot2 was present in those from the phot1 phot2 double mutant. However, the same amounts of plasma membrane H+-ATPase were found in all of these plants. H+ pumping was induced by blue light in isolated guard cell protoplasts from the wild type, from the single mutants of phototropins (phot1-5 and phot2-1), and from the zeaxanthin-less mutant (npq1-2), but not from the phot1 phot2 double mutant. Moreover, increased ATP hydrolysis and the binding of 14-3-3 protein to the H+-ATPase were found in response to blue light in guard cell protoplasts from the wild type, but not from the phot1 phot2 double mutant. These results indicate that phot1 and phot2 mediate blue light-dependent activation of the plasma membrane H+-ATPase and illustrate that Arabidopsis guard cell protoplasts can be useful for biochemical analysis of stomatal functions. We determined isogenes of the plasma membrane H+-ATPase and found the expression of all isogenes of functional plasma membrane H+-ATPases (AHA1-11) in guard cell protoplasts.

In Bhutan, weedy rice (Oryza sativa L.) was grown together with cultivated rice on terraced paddy fields lower than 2620 m above sea level. Seeds of cultivars and weedy strains were collected at 22 collection sites located from 1000 to 2620 m above sea level. Cultivars with round seeds were frequently found in fields higher than 2250 m, and those with slender seeds in fields lower than 1630 m. All cultivars and weedy strains were divided into indica or japonica types by isozyme (multi-locus) and morpho-physiological (multicharacter) analyses. Japonica cultivars predominated in highland; Indica cultivars predominated in lowland. Plastid type was confirmed by the length polymorphism for the ORF100 region. The japonica cultivars carried non-deletion type ORF100. The indica cultivars carried deletion type ORF100. In contrast, weedy strains showed discrepancy in the combination of the nuclear and cytoplasm types. An intermediate type was found in weedy strains for isozyme genotypes. A recombinant type, which has indica genotypes for isozyme analysis with japonica cytoplasm, and vice versa, was frequently found in weedy strains. These findings suggested that weedy strains would be generated by natural hybridization between indica and japonica. Further, they might fail to recombine their prior genotypes sets for isozyme and cytoplasm. Morphophysiological characters did not show such a tendency. Morpho-physiological analysis of highland plants, in particular, showed indica cultivars and weedy strains with japonica cytoplasm. Hypervariable nuclear microsatellite analysis was then used to compare cultivars and weedy strains. Identical alleles were shared between indica and japonica, and also between cultivars and weedy strains. This suggested that there was gene-flow resulting from natural hybridization.

Our results suggest that PLASMA sterilization may be unsuccessful under certain conditions, particularly when used for items with complex shapes and narrow lumens. Alternatively, LTSF sterilization demonstrates excellent efficacy and is comparable to EOG sterilization. LTSF could potentially act as a substitute if EOG becomes unavailable due to environmental concerns.

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