Effect of ABA,arginine and sucrose on protein content of date palm somatic embryos

Abscisic acid (ABA), arginine and sucrose were evaluated for their effects on the morphology, germination rates and protein content of date palm somatic embryos (SE). Different concentrations of these supplements in the culture medium were used. The comparative study of SE length and thickness between treated and untreated SE revealed no differences, except for ABA (20 μM), which increased thickness. A decrease of water content (WC) in favor of an increase in dry weight (DW) was observed in all treated SE, especially with sucrose (90 g l⁻¹) and ABA (20 μM). Only ABA (20 and 4 μM) caused a proliferation rate of the cultures higher than those in the control. Although all the tested compounds increased protein content, ABA (20 μM) was more effective in protein enrichment than arginine and sucrose treatments. The SDS-PAGE protein profiles showed a significant difference between treated and untreated SE. A protein band of 22 kDa, identified as glutelin in a previous work, was accumulated after treatment with 20 μM ABA or 3 mM arginine. These findings may contribute to further understanding of the mechanisms involved in the accumulation of specific storage proteins in several plants.     

二硫苏糖醇对刺参卵母细胞体外成熟的影响

用海星鞘神经提取液和二硫苏糖醇对刺参卵母细胞进行了体外人工促熟研究。结果显示,二硫苏糖醇对刺参卵母细胞具有明显的促熟作用。刺参卵母细胞包被在滤泡中发育,卵母细胞通过动物极的卵突起连接到单层细胞构成滤泡膜上,充分发育的卵母细胞处于生发泡期,染色质凝集在核仁周围,处于生发泡期的卵母细胞在海水中不会自发成熟。用二硫苏糖醇处理充分发育的卵母细胞20min,卵母细胞的生发泡开始移动,并锚定在卵母细胞的动物极,继而出现生发泡破裂。此后,减数分裂重新启动,第一、二极体分别排出。但海星鞘神经提取液对刺参卵母细胞没有促熟作用。实验表明,二硫苏糖醇浓度在10-5～10-1mol/L范围内,均可促使刺参卵母细胞发生生发泡破裂,10-2mol/L时诱导卵母细胞成熟率可达90%以上。     

Characterization of wheat gluten subunits by liquid chromatography – Mass spectrometry and their relationship to technological quality of wheat

The quality of common wheat is largely influenced by the composition of its storage proteins. The currently presented research explores factors influencing observed differences in quality and quantity between wheat cultivars, in particular in relation to gluten composition and its relationship to technological characteristics. Eight wheat cultivars (H. Wieser, Seilmeier, W., Belitz, H.D., 1994 Parsi, Sirvan, Sivand, Pishgam, Pishtaz)were selected for evaluation. Analysis results demonstrated that Morvarid and Sirvan cultivars yielded the highest quality of wheat, while the Chamran cultivar was indicated as the most favorable for baking Taftoon bread. Conversely, the Sepahan cultivar was deemed to have the worse quality in both categories. A Q Exactive LC-MS/MS system was employed to evaluate the most effective sub-fractions of gliadin and glutenin on wheat quality. Matching peptides resulting from trypsin digestion on gliadin and glutenin fractions, led to the identification of subunits α/β-gliadin, γ-gliadin, HMW-Dx5, HMW-Bx17, HMW-Dy3, HMW-Dy10, HMW-By15, LMW-m, LMW-s, and LMW-i. The obtained results indicated that the most influential subunits of glutenin on wheat quality were Dy10, Dy3 and Dx5, while the most effective gliadin subfraction was noted to be α/β-gliadin However, the most important subunit influencing the quality of flat breads in particular was identified as the x-HMW-GS, in particular the Bx17 subunit, and LMW-GS.     

Quantitative LC-MS proteoform profiling of intact wheat glutenin subunits

Methodology for the quantitative analysis of intact glutenin proteins by ESI-LC-MS was developed with 1 Da mass resolution, constituting the first published application of proteoform profiling to plant biology. Two parent lines and 28 F5 crosses were analyzed in two blocks, 14 months apart. Control sample data were used to align retention times and normalize abundance between sample sets. A total of 4622 observations of 347 distinct proteoforms between 17 899 and 88 744 Da were observed. Proteoform abundances spanned a 1000-fold range and were linear (r² > 0.990) with dilution. A novel method for the objective elimination of low intensity, noise-dominated data using replicate variability within the dataset is presented. Two abundant PTMs were detected; one known but uncharacterized Bx and Dy high-molecular-weight glutenin subunit (HMWGS) PTM and the other in 24 low molecular weight proteoform pairs. Finally, 16 abundant proteoforms were detected in progeny but not in either parent. This application should increase the statistical power of correlations between gluten complement and functional data and drive the detection of novel PTMs that may indicate differential regulation of the cellular processes related to quality.     

Changes in carbohydrate levels and their metabolic enzymes in leaves,phloem sap and mesocarp during cucumber (Cucumis sativus L.) fruit development

Levels of carbohydrates and activities of metabolic enzymes were examined in leaves (source), phloem sap (flow) and mesocarp tissues (sink) in the course of cucumber (Cucumis sativus L.) fruit development, from 2 days before anthesis to 20 days after anthesis. While total sugar levels increased in all the three sampling organs, starch levels declined in leaves and mesocarp tissues as fruit development progressed. Glucose and fructose were the primary contributors to the soluble sugar pools in mature leaves. Stachyose was found as the most important component of the phloem sap extracts, followed by sucrose and raffinose. However, the primary sugars accumulated in mesocarp tissues were glucose and fructose, not stachyose or sucrose. Activities of sucrose synthesizing enzymes (sucrose phosphate synthase plus sucrose synthase in the synthesizing direction) exceeded that of sucrose degrading enzymes (acid invertase, neutral invertase plus sucrose synthase in the degrading direction) in leaves, which might cause a sucrose pool utilized in raffinose and stachyose biosynthesis. While alkaline a-galactosidase form I activity declined, stachyose synthase activity showed a rapid increase until 12 days after anthesis and only subsequently decreased in leaves. Activities of sucrose degrading enzymes were always much higher than that of sucrose synthesizing enzymes in mesocarp tissues. Thus, sucrose accumulation could not occur in mesocarp tissues. While stachyose synthase activity steadily decreased, alkaline a-galactosidase form I activity showed a moderate increase before decrease in mesocarp tissues. The relationship between levels of soluble sugars and activities of relative enzymes was also discussed.     

牛精子原位杂交DTT去凝集条件优化

[目的]确定牛精子原位杂交DTT去凝集处理的优化浓度及时同.[方法]牛精子解冻后经密度梯度离心优选并制片,采用0、5、10、20、40、80和100 mmol/L六个DTT浓度以及15、30、45和60 min4个时间段的28个组合进行去凝集处理,Motic Images Advanced 3.2软件测量精子解聚后头部面积.[结果]40 mmol/L DTT处理45min精子头部面积为(52.32±4.68 )μm2,浓度＜ 40 mmol/L的16个处理组及浓度≥40 mmol/L、处理时间＜45 min的6个处理组,精子头部面积小于(52.32±4.68) μm2(p＜ 0.01);浓度＞40 mmol/L、时间≥45 min的5个处理组,精子头部面积与40 mmol/L DTT 45 min组差异不显著(P＞0.05),但精子形态不完整.[结论]40mmol/L DTT 45 min为较优处理组合.     

Expression,subcellular localization and phytohormone stimulation of a functional sucrose transporter (MdSUT1) in apple fruit

Sucrose transporters or carriers (SUTs or SUCs) play a crucial role in the cell-to-cell distribution of sucrose throughout the plant. Our data demonstrated that MdSUT1 is expressed almost ubiquitously in leaves, shoots, flowers and fruits, and the MdSUT1 was localized in plasma membranes of both sieve element/companion cell complex and storage parenchyma cells in apple fruits detected by immuno-gold labeling. The MdSUT1 expression was rapidly up-regulated by abscisic acid (ABA) at the translational levels in the treatments of the fruit tissues. Ten μmol L⁻¹ ABA and gibberellin acid (GA) induced MdSUT1 protein amounts dramatically, whereas 6-benzyl aminopurine (6-BA), indole-3-acetic acid (IAA) and cis-(−) ABA showed no distinct induction effect. These results suggest that MdSUT1 may be a component of ABA signaling pathways involved in the regulation of photoassimilate transport. ABA could function in plant sucrose trans-membrane transport, thus providing a clue that may help to unravel cross-talk between plant hormone and sucrose signaling pathway.     

Extractability and chromatographic separation of rye (Secale cereale L.) flour proteins

A size exclusion – high performance liquid chromatography (SE-HPLC) method originally developed for separating wheat, barley or rice proteins was applied to study the extractability and molecular weight (MW) distribution of rye flour proteins. These were extracted with 50 mmol/l sodium phosphate buffer (pH 6.8) containing 2.0% (w/v) sodium dodecyl sulfate (SDS) and, optionally, 1.0% (w/v) dithiothreitol (DTT). About 95% of the proteins were extracted in buffer containing 2.0% SDS. Addition of 1.0% DTT to such buffer increased the protein extractability to 100%, indicating that rye flour contains some proteins cross-linked by disulfide (SS) bonds. The SE-HPLC profiles revealed that rye flour contains SS-linked HMW-secalins and 75 k γ-secalins which elute in specific peaks. Upon reduction, these SS-linked protein aggregates dissociate and some entrapped albumins, globulins and/or ω-secalins are released. Rye flour albumins and globulins elute over the entire SE-HPLC profile. In contrast, the monomeric ω-secalins and 40 k γ-secalins are detected in specific well resolved SE-HPLC peaks. The applied fast and reproducible method can be used to characterise and quantify rye flour proteins and to determine changes as a result of processing.     

Is DTT a means to improve the mutation spectrum of vegetatively propagated plants?

Summary Freshly detached leaves of Achimenes cv. Cupido were placed, with their petioles, into a solution of DTT (dithiothreitol) during one or more hours, prior to irradiation with X-rays or fast neutrons. The radioresistance increased considerably, requiring a doubling of the X-ray dose to obtain the same survival and production of rhizomes as with non-pretreated leaves. The protection against fast neutrons was less. The mutation frequency decreased after pretreatment with DTT, though more drastically with X-rays than with fast neutrons. It seems that, in comparison with non-pretreatment of leaves the relative number of drastic mutants (supposedly caused by gross chromosome aberrations) hardly deviates significantly when leaves are pretreated with DTT, both after X-rays and fast neutrons; i.e. no improvement of the mutation spectrum has been obtained.     

Magnesium deficiency induced oxidative stress and antioxidant responses in mulberry plants

The aim of the study was to implicate induction of oxidative stress and antioxidative responses with the effects of Mg deficiency in mulberry plants. Mulberry (Morus alba L.) cv. Kanva-2 plants grown in hydroponics were subjected to deficiency of Mg. Mg-deficient plants developed visible symptoms—deep interveinal chlorotic mottling and necrosis in the older and middle leaves. The decreases in the dry matter yield of plants and concentrations of sugars and starch in the leaves of Mg-deficient plants are suggestive of decreased photosynthetic activity. Mg-deficiency decreased concentrations of photosynthetic pigments, and increased concentrations of H₂O₂ and ascorbate and activities of antioxidative enzymes—peroxidase (POD), ascorbate peroxidase (APX) and superoxide dismutase (SOD). The results suggest induction of oxidative stress by enhancing generation of ROS and inducing alterations in redox status, accompanied by activation of antioxidant machinery including induction of some new SOD isoforms in Mg-deficient mulberry plants. Despite significant increase in H₂O₂, lipid peroxidation was decreased in Mg-deficient plants.