An efficient protocol of callus induction, plant regeneration and long-term maintenance of embryogenic cultures for manilagrass was developed. Callus induction and embryogenic callus formation were influenced by cytokinins and nodal positions. Murashige and Skoog (MS) medium with 2 mg l -1 2,4-dichlorophenoxyacetic acid (2,4-D), 0.02 mg l -1 kinetin (KT) or 6-benzyladenine (BA) gave the highest frequency for both callus induction and embryogenic callus formation compared with 0.02 mg l -1 thidiazuron (TDZ) or N 6 -(2-isopenteny) adenine (2iP). The frequency of callus induction of different nodes (from the first to the sixth node) varied from 22.5 to 92.1%, and the embryogenic callus formation frequencies ranged from 13.3 to 25.7%. The highest frequencies of callus induction and embryogenic callus formation (92.1 and 25.7%, respectively) were observed in the fourth node group. During subculture on callus induction and maintenance medium, somatic embryos formed on the surface of the embryogenic callus. On regeneration medium, the regeneration rates of embryogenic callus varied from 96.8 to 100% during the 4-year period of subculture. The results also indicate that preservation of manilagrass callus is stable at low-temperature (4°C) over a period of 11 months. No significant differences were found in the activities of superoxide dismutase (SOD), peroxidase (POD) and proline content of the plants regenerated from the 4-year subcultured callus on different regeneration media.

To study growth in the presence of NaCl, in vitro plantlets regenerated from callus of manilagrass (Zoysia matrella [L.] Merr.) were cultured on regeneration medium supplemented with or without 0.3 M NaCl. The results indicated that growth was significantly inhibited by NaCl, with the leaves becoming relatively shorter and thicker. The differences of in vitro plantlets grown under NaCl stress provided specific criteria for the selection of salt tolerant variants. The 6-year maintained calli were treated with different doses (0, 5, 10, 20, 40, 80, 100, 150, 200, 250, and 300 Gy) of 60 Co c rays. Regeneration rate and regeneration capacity of the calli were highest after treatment with 20 Gy 60 Co c rays, 27.08 and 91.67% respectively. When the irradiation dose was increased to 100 Gy, 10.42% of the calli developed shoots, but at 150 Gy, both regeneration capacity and regeneration rate declined significantly, and no shoot was observed after 6 weeks of regeneration. Therefore, 100-150 Gy is the most appropriate irradiation span for inducing somaclonal variation. The irradiated calli were selected in vitro for NaCl tolerance. Five NaCl tolerant variant lines, Ze1, Fv1, Te1, Tw1, Fr1, were selected on subculture medium supplemented with 0.35 M NaCl, then transferred to regeneration medium containing 0.25 M NaCl, and grown in a greenhouse. The dark green colour index (DGCI) was used to identify the amount of injury caused by NaCl treatment. This was significantly higher in four of the lines, Ze1, Fv1, Te1, Fr1 (30.88, 31.17, 30.45 and 37.70%, respectively) compared to the control line (CK), which was regenerated from calli subcultured monthly (27.39%), indicating that watering with NaCl caused less injury in these four lines. These lines had lower proline contents than CK under salt stress. The superoxide dismutase (SOD) activity was higher in Ze1 under control condition and its peroxidase (POD) activity increased significantly under salt stress. With Fr1 catalase (CAT) activity was higher under salt stress. The higher activity of these antioxidant enzymes may contribute to the enhanced salt tolerance of the four plant lines.

Lead tolerance and accumulation in shoots and edible parts varied with crop species and soil type. The critical Pb concentrations at 10% yield reduction were 24.71, 28.25, and 0.567 mg kg(-1) for pakchoi, celery, and hot pepper, respectively under hydroponic conditions, whereas were 13.1, 3.83, 0.734 mg kg(-1) grown in the Inceptisol and 31.7, 30.0, 0.854 mg kg(-1) in the Alluvial soil, respectively. Based on the threshold of human dietary toxicity for Pb, the critical levels of soil available Pb for pakchoi, celery, and hot pepper were 5.07, 8.06, and 0.48 mg kg(-1) for the Inceptisol, and 1.38, 1.47, and 0.162 mg kg(-1) for the Alluvial soil, respectively. Similarly, the total soil Pb thresholds were different from vegetable species and soil types.

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Background Plant height, as one of the important agronomic traits of rice, is closely related to yield. In recent years, plant height-related genes have been characterized and identified, among which the D3 gene is one of the target genes of miR528, and regulates rice plant height and tillering mainly by affecting strigolactone (SL) signal transduction. However, it remains unknown whether the miR528 and D3 interaction functions in controlling plant height, and the underlying regulatory mechanism in rice. Results In this study, we found that the plant height, internode length, and cell length of internodes of d3 mutants and miR528-overexpressing (OE-miR528) lines were greatly shorter than WT, D3-overexpressing (OE-D3), and miR528 target mimicry (OE-MIM528) transgenic plants. Knockout of D3 gene (d3 mutants) or miR528-overexpressing (OE-miR528) triggers a substantial reduction of gibberellin (GA) content, but a significant increase of abscisic acid (ABA) accumulation than in WT. The d3 and OE-miR528 transgenic plants were much more sensitive to GA, but less sensitive to ABA than WT. Moreover, the expression level of GA biosynthesis-related key genes, including OsCPS1, OsCPS2, OsKO2 and OsKAO was remarkably higher in OE-D3 plants, while the NECD2 expression, a key gene involved in ABA biosynthesis, was significantly higher in d3 mutants than in WT and OE-D3 plants. Conclusion The results indicate that the miR528-D3 module negatively regulates plant height in rice by modulating the GA and ABA homeostasis, thereby further affecting the elongation of internodes, and resulting in lower plant height, which adds a new regulatory role to the D3-mediated plant height controlling in rice.