Ja Ock Guh

Generation of Resistance to the Diphenyl Ether Herbicide, Oxyfluorfen, via Expression of the Bacillus subtilis Protoporphyrinogen Oxidase Gene in Transgenic Tobacco Plants.

In an effort to develop transgenic plants resistant to diphenyl ether herbicides, we introduced the protoporphyrinogen oxidase (EC 1.3.3.4) gene of Bacillus subtilis into tobacco plants. The results from a Northern analysis and leaf disc assay indicate that the expression of the B. subtilis protoporphyrinogen oxidase gene under the cauliflower mosaic virus 35S promoter generated resistance to the diphenyl ether herbicide, oxyfluorfen, in transgenic tobacco plants.

Characterization of a glutathione S-transferase gene ATGST 1 in Arabidopsis thaliana

Abstract A glutathione S-transferase (GST) gene was cloned in Arabidopsis thaliana. The gene, designated ATGST 1, contained the entire transcription unit in three exons interrupted by two introns. The combined sequence of three exons had an open reading frame which predicted a GST protein of 208 amino acids. Gene transcription has been reported to be induced by pathogen attack and dehydration. In the present study northern blot analysis using a gene-specific DNA probe in the 3′ untranslated region revealed that expression of the gene was also rapidly induced by other environmental stresses such as wounding, low temperature, high salt and DPE herbicide treatment. The promoter region of the gene contained the sequence motif ATTTCAAA that is known to be present in ethylene-responsive elements and other motifs that are highly conserved amongst stress-inducible gene promoters.

Rapid diagnosis of resistance to sulfonylurea herbicides in monochoria (Monochoria vaginalis)

Abstract Sulfonylurea (SU)-resistant monochoria has recently been found in rice paddies in Korea. A quick and accurate means of confirming herbicide resistance is necessary to take timely management decisions. This article describes a rapid and reliable assay to detect SU-resistant biotype of monochoria. The techniques tested include seed germination, in vivo and in vitro acetolactate synthase (ALS; EC 4.1.3.18) activity, leaf, and whole-plant bioassays. In the whole-plant bioassay, shoot dry weight of the resistant (R) biotype was 3,200-fold less affected by imazosulfuron and sevenfold less affected by pyrazosulfuron-ethyl than the susceptible (S) biotype. Although the whole-plant bioassay is reliable, it is expensive, requires a lot of infrastructure, and takes a few months to complete. The germination rate of the R biotype in petri dish bioassays was > 200-fold less inhibited by imazosulfuron and 100-fold less inhibited by pyrazosulfuron-ethyl than that of the S biotype. Seed germination bioassays in petri dishes do not require as much infrastructure as whole-plant bioassays do and can be completed in a shorter time. Leaf bioassays showed that leaf color of the R biotype was > 1,600- and 300-fold less affected by imazosulfuron and pyrazosulfuron-ethyl, respectively, compared with that of the S biotype. This assay takes about 6 d to complete. In vivo ALS assays showed lower levels of resistance to ALS herbicides than did in vitro ALS assays, where the R biotype was about 200- and 30-fold less sensitive to imazosulfuron and pyrazosulfuron-ethyl, respectively, than the S biotype. All assays successfully distinguished the R from the S biotype, but in vitro ALS assays are the simplest and the quickest. The in vitro ALS assay was chosen as the standard procedure for future confirmation of resistance in monochoria populations. Caution is needed because the in vitro assay is not appropriate in cases wherein the resistance mechanism is increased metabolism of the herbicide or overexpression of the target enzyme. Results should be interpreted in relation to field history and field observations. Follow-up studies also are needed to verify that other resistance mechanisms do not confound the in vitro assay. Nomenclature: Imazosulfuron; pyrazonsulfuron-ethyl; monochoria, Monochoria vaginalis (Burm. f.) Kunth MOOVA; rice, Oryza sativa L.

Mechanism of paraquat tolerance in cucumber leaves of various ages

Abstract Differential sensitivity to paraquat was observed between cucumber cultivars and leaf age. Physiological responses to paraquat, including antioxidative enzyme activity, were investigated in leaf age classes of cucumber to identify mechanisms of paraquat tolerance. Leaf injury for ‘Naeseosamcheok’, ‘Daehandadagi’, ‘Baekgwangdadagi’, ‘Sangrokheukjinju’, and ‘Eunseongbaekdadagi’ cultivars was less than that of six other cultivars tested, averaged over leaf age and herbicide rate. The level of foliar injury caused by paraquat was Leaf 1 > 2 > 3 > 4 in seven of 11 cultivars used, where 4 was the youngest leaf. There was a positive correlation between leaf age and its relative susceptibility to paraquat, regardless of growth stage. Lipid peroxidation was less in the youngest leaf (Leaf 4) than in the older leaves at all herbicide concentrations. The youngest leaf had higher values for apparent photosynthesis than the oldest leaf. Differential leaf response to paraquat was partially correlated with the change in superoxide dismutase, ascorbate peroxidant, and glutathione reductase activities in treated leaves. Enzyme activity of ascorbate peroxidase (APX) was higher in Leaf 4 than in Leaves 1, 2, or 3 in untreated plants and after exposure to paraquat. APX isozymes were more abundant in treated than in untreated leaves and produced in higher amounts in younger than in older leaves. Application of ascorbate and glutathione before paraquat treatment protected cucumber leaves from paraquat injury. Nomenclature: Paraquat; cucumber, Cucumis sativus L.

Either Soluble or Plastidic Expression of Recombinant Protoporphyrinogen Oxidase Modulates Tetrapyrrole Biosynthesis and Photosynthetic Efficiency in Transgenic Rice

Protoporphyrinogen oxidase (Protox) is the last shared enzyme of the porphyrin pathway. As a continuation of our previous work in which the transgenic rice plants expressing the Bacillus subtilis Protox in the cytoplasm or the plastid showed resistance to diphenyl ether herbicide, this study was undertaken to identify the effects of tertapyrrole biosynthesis in these transgenic rice plants. The transgenic plants either targeted into plastids or expressed in cytoplasm showed higher Protox activity than wild-type plants did. Photosynthetic activity, measured as a quantum yield of photosystem II, was slightly higher in transgenic plants than in wild-type plants, but chlorophyll contents were not significantly different between transgenic and wild-type plants. As for porphyrin biosynthesis, both cytoplasm-expressed and plastid-targeted transgenic plants showed increased synthesis of aminolevulinic acid, Mg-Proto IX, and protoheme in comparison to wild-type plants whereas synthesis of protoporphyrin IX was similar for wild-type and transgenic plants. These results indicate that either cytoplasm or plastid expression of B. subtilis Protox in rice can upregulate the porphyrin pathway leading to increase in photosynthetic efficiency in plants.

Expression of recombinant protoporphyrinogen oxidase influences growth and morphological characteristics in transgenic rice

Transgenic rice plants expressing a Bacillus subtilis protoporphyrinogen oxidase (Protox), the last shared enzyme of the porphyrin pathway, in the cytoplasm (C89) or the plastids (P72) were compared with wild-type rice plants in their growth characteristics. Production of tiller buds 18 d after seeding was more profuse in transgenic plants than in wild-type plants, especially in plastid-targeted plants. Transgenic plants had 12–27% increase in tiller number and 17–33% increase in above-ground biomass compared with wild-type plants 4 and 8 weeks after transplanting of 2-week-old rice seedlings, demonstrating that tiller production and above-ground biomass correlate with each other. Cytoplasm-expressed and plastid-targeted transgenic plants also had a distinct phenotypic characteristic of narrower and more horizontal leaves than wild-type plants. Phenotypic and anatomical characteristics of the transgenic plants were clearly different from wild-type plants, indicating that regulation of porphyrin biosynthesis by expression of B. subtilis Protox in rice influences morphological characteristics of plant growth as well as biomass.

NUTRIENT DEPRIVATION AFFECTS XYLEM SAP FLOW AND WATER CHANNEL FUNCTION IN TOMATO PLANTS

ABSTRACT Heat-balance sap-flow gauge was used to measure xylem sap flow through the main stem of tomato (Lycopersicum esculentum Mill. cv. Young-Kwang) plants grown under nitrogen- or phosphorus-deficient nutrient solutions for nine days. Nitrogen deficient plants transported less sap than phosphorus deficient plants. Moreover, the effect of nitrogen deficiency on xylem sap flow was rapid, appearing within one day of treatment. Addition of mercury to nutrient solution induced immediate and drastic reduction in sap flow of both control and nutrient deprived plants. Subsequent use of dithiothreitol (DTT), however, did not recover sap flow in nutrient deprived plants. This suggests that the alteration in the characteristics of water channel was due to nutrient deficiencies.Heat-balance sap-flow gauge was used to measure xylem sap flow through the main stem of tomato (Lycopersicum esculentum Mill. cv. Young-Kwang) plants grown under nitrogen- or phosphorus-deficient nutrient solutions for nine days. Nitrogen deficient plants transported less sap than phosphorus deficient plants. Moreover, the effect of nitrogen deficiency on xylem sap flow was rapid, appearing within one day of treatment. Addition of mercury to nutrient solution induced immediate and drastic reduction in sap flow of both control and nutrient deprived plants. Subsequent use of dithiothreitol (DTT), however, did not recover sap flow in nutrient deprived plants. This suggests that the alteration in the characteristics of water channel was due to nutrient deficiencies.