The objectives of this study were to examine antioxidant enzyme responses to drought stress and rewatering at both enzymatic activity and transcript levels and to determine the major antioxidant processes associated with drought tolerance and post-drought recovery for a perennial grass species, kentucky bluegrass (Poa pratensis). Antioxidant enzyme responses to drought and rewatering in a drought-tolerant cultivar (Midnight) and a drought-sensitive cultivar (Brilliant) were compared in a growth chamber. Plants were exposed to 22 days of drought stress for ‘Midnight’ and 18 days for ‘Brilliant’ before rewatering to allow the leaf relative water content (RWC) of both cultivars to drop to the same level. ‘Midnight’ exhibited higher photochemical efficiency (Fv/Fm) and lower electrolyte leakage compared with ‘Brilliant’ when at the same water deficit status (26% to 28% RWC). After 6 days of rewatering, all physiological parameters returned to the control level for ‘Midnight’, but only Fv/Fm fully recovered for ‘Brilliant’. The transcript level of cytosolic copper/zinc superoxide dismutase (cyt Cu/Zn SOD) and ascorbate peroxidase (APX) was significantly higher in ‘Midnight’ than in ‘Brilliant’ when exposed to the same level of water deficit (26% to 28% RWC), suggesting that SOD and APX could be involved in scavenging oxidative stress-induced reactive oxygen species in kentucky bluegrass through changes in the level of gene expression. Significantly higher activities of APX, monodehydroascorbate reductase, glutathione reductase, and dehydroascorbate reductase as well as lower lipid peroxidation levels were observed in ‘Midnight’ versus ‘Brilliant’ when exposed to drought. However, the activities of SOD, catalase (CAT), and guaiacol peroxidase (POD) did not differ between the two cultivars. After 6 days of rewatering, ‘Midnight’ displayed significantly higher activity levels of CAT, POD, and APX compared with ‘Brilliant’. The enzyme activity results indicate that enzymes involved in the ascorbate–glutathine cycle may play important roles in antioxidant protection to drought damage, whereas CAT, POD, and APX could be associated with better post-drought recovery in kentucky bluegrass.
Lixin Xu, Liebao Han, and Bingru Huang
Li Xu, Suzhen Huang, Yulin Han, and Haiyan Yuan
An improved three-stage protocol for plant regeneration via somatic embryogenesis of the horticulturally important plant Iris germanica L. was developed using shoot apex segments as explants. At the first stage of the experiment, 60% of callus was obtained from shoot apex segments of I. germanica on Murashige and Skoog’s (MS) medium supplemented with 4.52 μm 2,4-dichloropheoxyacetic acid (2,4-D) and 0.44 μm 6-benzyladenine (6-BA). When nonembryogenic calli were subcultured on MS medium with 11.31 μm 2,4-D and 0.44 μm 6-BA, maximum frequency of embryogenic callus (66.0%) was obtained. At the second stage, the treatment of 9% (w/v) sucrose resulted in the optimum somatic embryo (SE) formation (70.0%). More than 90.0% of SEs germinated with bipolar structure and regenerated into plantlets on plant growth regulator-(PGR)free MS medium during the third stage. Regenerated plantlets were successfully acclimatized in greenhouse environment with little somaclonal variation. Histological study showed that somatic embryogenesis stages were asynchronous and SEs developed from the surface and inner tissue of embryogenic calli.
Lixin Xu, Mili Zhang, Xunzhong Zhang, and Lie-Bao Han
Zoysiagrass (Zoysia spp.), a warm-season turfgrass species, experiences freezing damage in many regions. The mechanisms of its cold acclimation and freezing tolerance have not been well understood. This study was designed to investigate effects of cold acclimation treatment on leaf abscisic acid (ABA), cytokinin (transzeatin riboside (t-ZR), and antioxidant metabolism associated with freezing tolerance in zoysiagrass (Zoysia japonica). ‘Chinese Common’ zoysiagrass was subjected to either cold acclimation treatment with temperature at 8/2 °C (day/night) and a photosynthetically active radiation (PAR) of 250 µmol·m−2·s−1 over a 10-hour photoperiod or normal environments (temperature at 28/24 °C (day/night), PAR at 400 µmol·m−2·s−1 and 14-hour photoperiod) for 21 days in growth chambers. Cold treatment caused cell membrane injury as indicated by increased leaf cell membrane electrolyte leakage (EL) and malondialdehyde (MDA) content after 7 days of cold treatment. Cold treatment increased leaf ABA and hydrogen peroxide content and reduced t-ZR content. Leaf superoxide dismutase (SOD), ascorbate peroxidase (APX) activity, and proline content increased, whereas catalase (CAT) and peroxidase (POD) activity declined in response to cold treatment. Cold treatment increased freezing tolerance as LT50 declined from −4.8 to −12.5 °C. The results of this study indicated that cold acclimation treatment might result in increases in ABA and H2O2, which induce antioxidant metabolism responses and improved freezing tolerance in zoysiagrass.
Na Zhang, Lu Han, Lixin Xu, and Xunzhong Zhang
Ethephon [ETH (2-chloroethylphosphonic acid, an ethylene-releasing compound)] has been used as a plant growth regulator in turfgrass management. The aim of the study was to assess the effects of ETH seed treatment on drought tolerance of kentucky bluegrass (Poa pratensis) seedlings. Seeds of two kentucky bluegrass cultivars, Midnight and Nuglade, were exposed to ETH treatment or untreated as controls. Seedlings were then exposed to two water regimes: well-watered conditions and polyethylene glycol (PEG)–induced drought conditions. ETH-treated plants exhibited better turf performance relative to the untreated control under PEG-stressed conditions illustrated by higher relative water content (RWC) and lower lipid peroxidation and lower electrolyte leakage (EL). In both cultivars, ETH treatment increased enzyme activity of ascorbate peroxidase (APX), peroxidase (POD), and catalase (CAT); proline content; and soluble protein content under PEG-induced drought conditions. The results suggest that ETH seed treatment can improve drought tolerance in kentucky bluegrass seedlings.
Feifei Li, Da Zhan, Lixin Xu, Liebao Han, and Xunzhong Zhang
Heat stress is a major limiting factor for growth of cool-season perennial grass species, and mechanisms of heat tolerance have not been well understood. This study was designed to investigate antioxidant enzyme and hormone metabolism responses to heat stress in two kentucky bluegrass (Poa pratensis L.) cultivars contrasting in heat tolerance. The plants were subjected to 20/20 °C [day/night (control)] or 38/30 °C [day/night (heat stress)] for 28 days in growth chambers. Heat stress increased leaf electrolyte leakage (EL) and malondialdehyde (MDA) with heat-tolerant cultivar EverGlade exhibiting lower levels of EL and MDA relative to heat-sensitive cultivar Kenblue under heat stress. Superoxide dismutase (SOD) and catalase (CAT) activity increased and then declined during 28 days of heat stress. Peroxidase (POD) and ascorbate peroxidase (APX) activity declined and then increased during heat stress. ‘EverGlade’ had greater activities of SOD, CAT, POD, and APX relative to ‘Kenblue’ under heat stress. In addition, ‘EverGlade’ had two additional SOD isozymes and three additional POD isozymes relative to ‘Kenblue’ under heat stress. Leaf abscisic acid (ABA) increased in response to heat stress. Leaf indole-3-acetic acid (IAA) increased and then declined during heat stress. ‘OverGlade’ had higher ABA and IAA content relative to ‘Kenblue’. At the end of heat stress, leaf IAA and ABA content were 27.8% and 73% higher in ‘EverGlade’ relative to ‘Kenblue’, respectively. The results indicated that antioxidant enzymes and the hormones (ABA and IAA) were associated with kentucky bluegrass heat tolerance. Selection and use of cultivars with higher IAA and ABA content and greater antioxidant enzyme activities may improve kentucky bluegrass growth and quality under heat stress.
Ruigang Wu, Yi Wang, Ting Wu, Xuefeng Xu, and Zhenhai Han
MYB (v-myb avian myeloblastosis viral oncogene homologs) transcription factors (TFs) are involved in diverse physiological processes, including cell shape determination, cell differentiation, and secondary metabolism, as well as abiotic stress response. In the present study, MdMYB4, an R2R3-MYB protein that is a homolog of Arabidopsis thaliana MYB4, was identified and characterized. Quantitative real-time polymerase chain reaction (qRT-PCR) expression analysis demonstrated that MdMYB4 is extensively expressed in various apple (Malus domestica) tissues and that its expression is induced by cold, osmotic, and salt stress. An MdMYB4-GFP fusion protein was localized in the nucleus of transformed onion (Allium cepa) epidermal cells and had a certain transcriptional activation activity by yeast one-hybrid assay. Overexpression of the MdMYB4 gene remarkably enhanced the tolerance of stably transgenic apple calli to severe salt and cold stress, and both the relative conductivity and malondialdehyde (MDA) accumulation of transgenic calli under salt and cold stress were significantly lower than in the wild type control. Taken together, these results suggest that MdMYB4 may play a positive regulatory role in both cold and salt stress responses.
Wanmei Jin, Jing Dong, Yuanlei Hu, Zhongping Lin, Xuefeng Xu, and Zhenhai Han
Dehydration response element binding (DREB)1b is a cold-inducible transcription factor in Arabidopsis thaliana. DREB1b driven by cauliflower mosaic virus 35S promoter was genetically introduced into grape Vitis vinifera L. cv. Centennial Seedless through Agrobacterium-mediated transformation for improving its cold resistance and exploring new genetic breeding approaches to obtain cold-resistant cultivars. In this study, Southern blot analysis showed the DREB1b gene was integrated into the transgenic grapevines with one to two copies. Northern blot analysis showed the presence of DREB1b transcripts in the independent transgenic lines 3, 5, 6, and 7. Further characterization of transgenic grapevines confirmed that both electrolyte leakage conductivity and the freezing point of the transgenic plants were lower than those of wild-type plants. After the cold treatment at –4 °C for 12 h, 26% of transgenic plants wilted among which 95% plants recovered once being placed under the condition of temperature 22 to 25 °C. However, subjected to the same treatment, 98% of nontransgenic plants wilted and only 2% recovered. Our results lead to the conclusion that activity of DREB1b in the transgenic grape could significantly improve its resistance to cold stress.
Haishan An, Feixiong Luo, Ting Wu, Yi Wang, Xuefeng Xu, Xinzhong Zhang, and Zhenhai Han
Fine root (≤2 mm in diameter) systems play a pivotal role in water and mineral uptake in higher plants. However, the impact of fine root architecture on tree growth and development is not fully understood, especially in apple trees. Here, we summarize a 6-year-trial study using minirhizotrons to investigate the relationships between fine root production, mortality, and longevity in ‘Red Fuji’ trees grafted on five different rootstocks/interstems. Based on root length density (RLD), fine root production and mortality were markedly lower in ‘Red Fuji’ trees growing on dwarfing M.9 (M.9) and Shao series no. 40 (SH.40) rootstocks than in trees on standard Malus robusta ‘Baleng Crab’ (BC) rootstock. The use of M.9 and SH.40 as interstems led to an extensive reduction in fine root production and mortality in comparison with BC rootstock. Root number density (RND), but not average root length (ARL), showed similar patterns to RLD. About one-half of fine roots in ‘Red Fuji’ tree growing on M.9 were scattered within the top 0–20 cm of topsoil, indicating shallow root system in M.9, whereas in trees on BC, 55.15% of fine roots were distributed between 100- and 150-cm soil depth, indicating a deep root architecture. The addition of interstems did not alter fine root soil-depth distribution. For all rootstocks/interstems, fine roots with a life span of less than 80 days were generated in spring and summer, but fine roots which lived for more than 81 days were produced almost all the year round. In conclusion, lower fine root numbers were associated with the dwarfing effect in dwarfing rootstocks/interstems, but ARL and shallower rooting were not.
Han Xu, Cuihua Bai, Wei Wang, Changmin Zhou, Luwei Zhu, and Lixian Yao
Free amino acid (FAA) profile is an important indicator of the quality of fruit and fruit product. Foliar nutrient diagnosis has been used for crop yield prediction for decades but not for fruit quality evaluation. Concentrations of 11 leaf nutrients including N, P, K, Ca, Mg, S, Fe, Mn, Cu, Zn, and B at stages of terminal shoot maturation and fruit development and fruit FAA profile at harvest were examined in longan in South China. The relation between leaf nutrient and fruit FAA was then investigated by multiple stepwise regression analysis. Foliar N content was greatest among the nutrients among the detected elements at both stages. Twenty-nine FAAs were determined in longan flesh, with alanine (19.9%), γ-aminobutyric acid (17.5%), glutamic acid (15.2%), and asparagine (10.7%) as the main components. Flesh individual FAA, essential amino acid (AA), umami-, and sweet and bitter taste AA strongly depended on foliar nutrients. However, the relation between flesh FAAs and foliar nutrients varied with FAA species. Leaf N was the dominant indicator for most pulp FAAs at two growth stages, while other nutrients (e.g., B, Zn, P, K, Ca, Mg) also played versatile roles on flesh FAAs. This work provides a novel tool to predict fruit FAAs via foliar nutrient diagnosis, which supports the practicality of producing specific target fruit or improving fruit quality through regulation of fertilization strategies in fruit production.
Li-Juan Zhang, Tian-Xiu Zhong, Li-Xin Xu, Lie-bao Han, and Xunzhong Zhang
Soil water deficit impacts cold acclimation and freezing tolerance in creeping bentgrass (Agrostis stolonifera L.), but the mechanisms underlying have not been well understood. The objectives of this study were to investigate the effects of deficit irrigation before and during cold acclimation on osmoprotectants, antioxidant metabolism, and freezing tolerance in creeping bentgrass. The grass was subjected to three-soil moisture levels: well-watered [100% container capacity (CC)], deficit irrigation induced-mild drought stress (60% CC), and severe drought stress (30% CC) for 35 days including 14 days at 24/20 °C (day/night) and then 21 days under cold acclimation treatment (2 °C) in growth chambers. Leaf proline and total soluble sugar (TSS) levels were higher in the grass under mild drought stress relative to that under severe drought stress. Superoxide (O2 −·), hydrogen peroxide (H2O2), and malondialdehyde (MDA) content were higher in the grass under severe drought relative to that under well-watered and mild drought stress at day 35. Mild drought stress increased catalase (CAT) and guaiacol peroxidase (POD) activity, induced new isoforms and increased band intensities of superoxide dismutase (SOD), CAT, and POD during cold acclimation (days 14 to 35). No differences in osmoprotectants, antioxidant metabolism, and freezing tolerance were found between mild drought and well-watered treatments. The results of this study suggest deficit irrigation-induced mild drought stress in late fall and winter could induce accumulation of osmoprotectants and improve antioxidant metabolism, and freezing tolerance, but severe drought stress could reduce freezing tolerance of creeping bentgrass in the region with limited precipitation.