Differential Interaction Between PAV-like Isolates of Barley Yellow Dwarf Virus and Barley (Hordeum vulgare L.) genotypes

Four PAV-like isolates of barley yellow dwarf virus (BYDV) were identified as causing very severe (RG), severe (2t), moderately severe (3b) and mild symptoms (13t) in barley (Hordeum vulgare) cultivar Plaisant in a growth chamber at 25 days after inoculation. These isolates had different effects on a range of barley genotypes. Cultivar Vixen, which contains the Yd2 resistance gene, and 80–81BQCB10 were not affected by any isolate. Five other genotypes were significantly affected by at least one of the isolates. Line Ea52 (which is a mutant of the Japanese cultivar Chikurine Ibaraki) was more susceptible to BYDV-PAV than Chikurin Ibaraki 1. No serological differences were detected between the four isolates using monoclonal or polyclonal antibodies. Virus antigen concentration, estimated by enzyme-linked immunosorbent assay (ELISA), was correlated with the decrease in the shoot fresh weight for all isolates and all genotypes except for Vixen and 80–81BQCB10. In field tests, the severity of symptoms induced by the BYDV-PAV isolates was in accordance with that estimated in the growth chamber. However isolate 2t was more severe on cultivar Vixen and overcame the partial resistance of Chikurin Ibaraki 1 to the three other isolates. The results show that virus antigen concentration not only contributes to characterizing the resistance levels of barley genotypes but also the severity of BYDV-PAV isolates.     

Differential Interaction Between PAV-like Isolates of Barley Yellow Dwarf Virus and Barley (Hordeum vulgare L.) genotypes

Four PAV-like isolates of barley yellow dwarf virus (BYDV) were identified as causing very severe (RG), severe (2t), moderately severe (3b) and mild symptoms (13t) in barley (Hordeum vulgare) cultivar Plaisant in a growth chamber at 25 days after inoculation. These isolates had different effects on a range of barley genotypes. Cultivar Vixen, which contains the Yd2 resistance gene, and 80-81 BQCB10 were not affected by any isolate. Five other genotypes were significantly affected by at, least one of the isolates. Line Ea52 (which is a mutant of the Japanese cultivar Chikurine Ibaraki) was more susceptible to BYDV-PAV than Chikurin Ibaraki 1. No serological differences were detected between the four isolates using monoclonal or polyclonal antibodies. Virus antigen concentration, estimated by enzyme-linked immunosorbent assay (ELISA), was correlated with the decrease in the shoot fresh weight for all isolates and all genotypes except for Vixen and 80-81BQCB10. In field tests, the severity of symptoms induced by the BYDV-PAV isolates was in accordance with that estimated in the growth chamber. However isolate 2t was more severe on cultivar Vixen and overcame the partial resistance of Chikurin Ibaraki 1 to the three other isolates. The results show that virus antigen concentration not only contributes to characterizing the resistance levels of barley genotypes but also the severity of BYDV-PAV isolates.     

Lysine Catabolism in Barley (Hordeum vulgare L.)

Lysine catabolism in seedlings of barley (Hordeum vulgare L. var. Emir) was studied by direct injection of the following tracers into the endosperm of the seedlings: aspartic acid-3-(14)C, 2-aminoadipic acid-1-(14)C, saccharopine-(14)C, 2,6-diaminopimelic acid-1-(7)-(14)C, and lysine-1-(14)C. Labeled saccharopine was formed only after the administration of either labeled 2,6-diaminopimelic acid or labeled lysine to the seedlings. The metabolic fate of the other tracers administered also supported a catabolic lysine pathway via saccharopine, and apparently proceeding by a reversal of some of the biosynthetic steps of the 2-aminoadipic acid pathway known from lysine biosynthesis in most fungi. Pipecolic acid seems not to be on the main pathway of l-lysine catabolism in barley seedlings.     

大麦黄花叶病严重度的遗传分析

本文就大麦黄花叶病(BYMV)的抗性进行了遗传分析。研究表明,在本研究中,大麦黄花叶病抗性表现为多基因控制的数量性状,符合“加性-显性”遗传模型,但主要受加性效应控制。回归分析与平均显性度((H_1/D)~(1/2))测定均表明为部分显性。且控制BYMV的严重度的显性基因数约为3—6组。遗传力估算较高。最后就实验结果对BYMV抗性育种进行了初步分析讨论。     

Phosphoinositides in Barley (Hordeum vulgare L.) Aleurone Tissue

[3H]Inositol labeling of barley (Hordeum vulgare L. cv Himalaya) aleurone layers and analysis of phospholipids by deacylation revealed the presence of phosphatidylinositol (PtdIns), PtdIns3P, and PtdIns4P but not PtdInsP2 species. In contrast to an earlier report (P.P.N. Murthy, G. Pliska-Matyshak, L.M. Keranen, P. Lam, H.H. Mueller, N. Bhuvarahamurthy [1992] Plant Physiol 98: 1498-1501) systematic chemical degradation of PtdIns revealed no evidence of a second isomer of PtdIns. Evidence of the widespread occurrence of 3-phosphorylated PtdIns within the plant kingdom is presented.     

Apoplast Acidification in Growing Barley (Hordeum vulgare L.) Leaves

Apoplast acidification associated with growth is well documented in roots, coleoptiles, and internodes but not in leaves. In the present study, advantage was taken of the high cuticle permeability in the elongation zone of barley leaves to measure apoplast pH and growth in response to application of test reagents. The role of the plasma membrane H⁺-ATPase (PM-H⁺-ATPase) and K⁺ in this process was of particular interest. pH microelectrodes and an in vitro gel system with bromocresol purple as pH indicator were used to monitor apoplast pH. Growth was measured in parallel or in separate experiments using a linear variable differential transformer. Test reagents that blocked (vanadate) or stimulated (fusicoccin) PM-H⁺-ATPase or that reduced (Cs⁺, tetraethylammonium) K⁺ uptake were applied. Apoplast pH was lower in growing than in nongrowing leaf tissue and increased in the elongation zone with increasing apoplast K⁺. Vanadate increased apoplast pH and reduced growth, whereas fusicoccin caused the opposite effects. It is concluded that barley leaves exhibit acid-growth-type mechanisms in that apoplast pH is lower in elongating leaf tissue. Both growth and apoplast pH depend on the activity of the PM-H⁺-ATPase and K⁺ transport processes. However, not all of the growth displayed by leaves is dependent on a lower apoplast pH in the elongation zone; up to 50 % of growth is retained when apoplast pH in the elongation zone increases to a value observed in mature tissue.     

大麦转基因座位结构的研究

利用质粒营救法获得基因枪法转化的4种转绿色荧光蛋白基因(green fluorescent protein,GFP)大麦的转基因座位序列,序列分析显示4种材料的转基因座位中均有完整栽体的串联重复现象,表明转基因整合是同源重组的结果.同时转基因座位中也存在不完整载体片段、基因组片段的混杂排列,说明转基因整合时也发生异常重组.微粒轰击的转基因整合是由异常重组和同源重组共同完成的.     

Flow Cytometric Analysis and Chromosome Sorting of Barley (Hordeum vulgare L.)

Flow cytometric analysis was systematically performed to optimize the concentration and duration of hydroxyurea (DNA synthesis inhibitor) and trifluralin (metaphase blocking reagent) treatments for synchronizing the cell cycle and accumulating metaphase chromosomes in barley root tips. A high metaphase index (76.5% in the root tip meristematic area) was routinely achieved. Seedlings of about 1.0-cm length were treated with 1.25 mM hydroxyurea for 14 h to synchronize the root tip meristem cells at the S/G2 phase. After rinsing with hydroxyurea, the seedlings were incubated in a hydroxyurea-free solution for 2 h and were treated with 1 microM trifluralin for 4 h to accumulate mitotic cells in the metaphase. The consistent high metaphase index depended on the uniform germination of seeds prior to treatment. High-quality and high-quantity isolated metaphase chromosomes were suitable for flow cytometric analysis and sorting. Flow karyotypes of barley chromosomes were established via univariate and bivariate analysis. A variation of flow karyotypes was detected among barley lines. Two single chromosome types were identified and sorted. Bivariate analysis showed no variation among barley individual chromosomes in AT and GC content.     

Effects of Red and Blue LED Light on the Growth and Photosynthesis of Barley (Hordeum vulgare L.) Seedlings

According to the action spectrum of photosynthesis, photosynthetic efficiency is highest for red light. However, long-term growth with only red light leads to unfavorable changes in plant morphology, decrease in photosynthetic capacity and plant productivity. Detailed mechanisms behind these changes are still poorly understood. We studied the effects of narrow-band red (RL) and blue (BL) LED lighting on the morphology and photosynthesis of barley (Hordeum vulgare L.) seedlings at 9 days old, when energy for plant growth comes mostly from the endosperm, and light has a mainly morphogenic effect on plant growth. Plants grown with white fluorescent lamps (WL) were used as a control. At this developmental stage, light spectrum had small but significant effects on most morphometric parameters, which may become more prominent as the plant grows. These effects were more pronounced in RL-grown plants and were similar to the ‘shade-avoidance response’, which is unusual as in nature it occurs when the fraction of red light in the spectrum is low. RL-grown plants also had impaired photosynthetic photochemical efficiency (as assessed by PAM-fluorometry and leaf absorption). BL-grown plants had a stronger similarity to control plants in their morphology and photosynthetic characteristics than RL-grown plants; however, they had higher NPQ and different NPQ induction kinetics than WL- and RL-grown plants. Our results suggest that photoregulation of plant morphology and photosynthesis evolutionarily adapted to natural light is miscoordinated in narrow-band LED light. We discuss possible reasons for this miscoordination and for the formation of observed phenotypes on the level of photoreceptors.

     

染色体重组对转基因大麦中gfp基因表达的作用

利用5种转绿色荧光蛋白基因(gfp)大麦不同株系及野生型大麦为材料,对不同转基因株系间以及转基因株系与野生型植株间进行杂交,分别对不同世代植株的根尖、花粉中gfp基因的表达量进行测定.结果表明,不同转基因株系间的根尖、花粉的gfp基因在表达量上存在差异,同一转基因材料的gfp基因表达存在组织差异;gfp基因在杂交后代中作为一个显性基因以孟德尔方式稳定遗传,不同染色体上的gfp基因重组有利于提高持基因表达.     

The Effect of Water Stress upon Polyamine Levels in Barley (Hordeum vulgare L.) Leaves

The effect of water stress on leaf polyamine content of fourHordeum vulgare varieties, Alger/Ceres, Palmella Blue, Rihaneand Roho, with different drought characters was studied After6 d without water Alger/Ceres, Palmella Blue and Rihane hadaccumulated putrescine, although only in concentrations up totwice those found in the controls, but Roho had a decreasedputrescine content. However, one common response was identified;the accumulation and subsequent loss of putrescine was dependentupon the maintenance and loss of leaf turgor respectively. Consequentlyvarietal differences in putrescine accumulation were relatedto water consumption rates and the extent of osmotic adjustment.Spermine behaved in a similar manner to putrescine but spermidinelevels always decreased. Polyamine levels were never high enoughto be an important component of solute accumulation. Prolinelevels were 150-fold higher and glycine-betaine levels 50-foldhigher than polyamines in stressed plants. Proline and glycine-betaineaccumulation occurred once a threshold turgor was reached, whichin Roho appeared to be a reduction by 0•2–0•25MPa or 30–40%. The importance of polyamine accumulationduring water stress is discussed. Key words: Hordeum vulgare, polyamine, water stress     

Effects of Plant Growth Regulators and Nutrient Supply on Tiller Bud Outgrowth in Barley (Hordeum distichum L.)

Seedlings of spring barley were raised in 100 and 20% nutrientsolution and treated with a foliar application of Terpal, Cerone,TIBA, GA₃ or BAP. The growth of individual tiller buds and tillers,the main shoot and the root system was recorded over the following15 d. Terpal and Cerone stimulated tiller bud elongation within5 d at both nutrient levels and after 15 d the number of emergedtillers was increased at the higher nutrient level. Terpal characteristicallypromoted the growth of secondary tiller buds whereas Ceronepromoted the emergence of the coleoptile tiller; both thesePGRs also retarded the development of the main shoot. TIBA increasedthe number of elongating tiller buds but this did not resultin greater tillering. GA₃ reduced the number of elongating tillerbuds and restricted their growth, especially in the high nutrientregime; this was accompanied by an increase in main shoot elongation.The growth and development of tiller buds was reduced by BAPand the number of emerged tillers was reduced at 15 d in bothnutrient levels; main shoot dry weight and root elongation werealso reduced. The results are considered in relation to theoverall influence of hormonal factors and mineral supply ontiller bud outgrowth. Hordeum distichum, spring barley, tiller bud outgrowth, plant growth regulators, Terpal, Cerone, GA₃, BAP, nutrient supply, apical dominance, TIBA     

Properties and Regulation of Aspartate Kinase from Barley Seedlings (Hordeum vulgare L.)

Aspartate kinase (EC 2.7.2.4) has been purified 8-fold and characterized from germinating barley (Hordeum vulgare) seedlings. The enzyme is inhibited 50% by 0.7 mm l-lysine and almost completely at 5 mm. l-Methionine does not affect the enzyme on its own, but at low concentrations (0.1-1 mm) increases the inhibition in the presence of lysine, indicating that the two amino acids act as cooperative feedback regulators.     

Proton Flux and Elongation in Primary Roots of Barley (Hordeum vulgare L.)

The elongation zone of the primary root of barley (Hordeum vulgare L.) has been reported to be markedly basic in pH, in apparent contradiction of the acid-growth theory. We determined simultaneously the location of the elongation zone and the basic zone in these roots and found them indeed to be the same. However, sections of barley root elongation zones were found to respond to acidic, basic, and neutral solutions as predicted by the acid-growth theory.     

The Uptake and Transport of Chromium by Barley Seedlings (Hordeum vulgare L.)

Uptake of ⁵¹CrO^2–₄ by intact barley seedlings was linearover 24 h and was stimulated by Ca2⁺ but inhibited by SO^2–₄and other Group VI anions. Uptake increased with increasingchromate concentration, but unless the concentration was high(100 µM) less than 1 per cent of the isotope absorbedwas transported to the shoots. The results of solvent extraction,subcellular fractionation and efflux studies indicated thatmost of the isotope accumulated by the roots was present ina soluble non-particulate form in the vacuoles. The possiblereasons for the restriction in chromate transport are discussedin relation to the metabolism of the element.     

Barley (Hordeum vulgare L.) inositol monophosphatase: gene structure and enzyme characteristics

The cellular myo-inositol (Ins) pool is important to many metabolic and signaling pathways in plants. Ins monophosphatase (IMPase; EC 3.1.3.25) activity is essential for the de novo synthesis of myo-Inositol (Ins), and for recycling of Ins in Ins(1,4,5)P3. However, proteins encoded by at least one family of IMP genes also have L-galactose-1-P phosphatase activity important to ascorbic acid synthesis, indicating a bifunctionality that links these two branches of carbon metabolism. As part of research into the regulation of Ins synthesis and supply during seed development, the barley IMP-1 gene and gene products were studied. The 1.4 kb barley IMP-1 promoter contains one low temperature response element (RE), two heat shock REs, one gibberellin and two auxin REs, and five sugar REs. Barley IMP-1 is expressed in all tissues assayed, and expression levels were not greatly altered by abiotic stress treatments. Reduced use of Ins for Ins P6 synthesis in developing seed of barley low phytic acid (lpa) mutants results in Ins accumulation, and IMP-1 expression is reduced in proportion to the increase in Ins level. The barley recombinant enzyme had a lower Km, indicating higher affinity, for D/L-Ins(3)P1 (Km = 9.7 microM) as compared with reported Km (Ins P1) values for other eukaryotic IMPases (43-330 microM) or with a reported Km (L-Gal-1P) of 150 microM for a kiwifruit (Actinidia deliciosa) enzyme. These and other data indicate that the barley IMP-1 gene is regulated at least in part in response to Ins metabolic needs, and that the enzyme it encodes displays catalytic properties well suited for a role in Ins synthesis, in addition to other roles as an L-gal-1-P phosphatase important to ascorbate synthesis, or as an IMPase important to Ins(1,4,5)P3 signal recycling.     

Proteins in Intercellular Washing Fluids from Leaves of Barley (Hordeum vulgare L.)

Primary leaves of barley were detached, infiltrated with variousbuffers, and centrifuged to yield ‘intercellular washingfluid’ (IWF). Effective pH control of the IWF was obtainedonly with Tris, among all buffers tried. In these liquids, upto 30 proteins were detected by gradient gel electrophoresis.Intracellular protein from injured cells at the cut ends ofleaves was present in IWF but did not contribute significantlyto the total protein recovered in this liquid. The yield ofprotein in the IWF depended on the buffer used for infiltrationand on the concentration of the buffer. Higher concentrationsof buffer yielded more protein. In other experiments leaves were infiltrated with Tris, centrifuged,and then infiltrated a second time with this buffer containingvarious concentrations of the zwitterionic detergent CHAPS,a sulphobetaine derivative of cholate. Gel electrophoresis ofthe IWF obtained after the second centrifugation revealed protein‘bands’ not detected when the detergent had beenomitted from the infiltration buffer. The electrophoretic patternsof protein ‘bands’ in the gels differed dependingon the CHAPS concentration used for infiltration. The effect of CHAPS on plasmalemma integrity was studied byobserving infiltrated tissue with the electron microscope andby treating isolated protoplasts with the detergent. After infiltrationwith CHAPS at 0.6 mM or 2.0 mM no plasmalemma breaks were detectedin leaves, and isolated protoplasts survived exposure to CHAPSat these concentrations for 2 h without bursting. Evidently,CHAPS at these low concentrations did not destroy the integrityof the plasmalemma; the additional protein recovered in theIWF under these conditions probably originated in the cell wall.Infiltration of leaves with 6.0 mM CHAPS resulted in breaksof the plasmalemma, in tissue collapse and leaf tip necrosis.Isolated protoplasts burst within minutes after being exposedto CHAPS at this concentration. Key words: Cell wall permeability, Intercellular space, Detergent, CHAPS, Protoplasts     

Mechanism of Septum Opening in Anthers of Two-rowed Barley (Hordeum vulgare L.)

To test the hypothesis that the rapid swelling of pollen grainsdriven by potassium movement opens the septum in anthers ofpoaceous plants, we studied (1) the behaviour of pollen grainsduring unfolding of the locule and (2) the distribution of potassiumin the locule in two-rowed barley. In the first experiment,the unfolding of decapitated anthers was observed. The pollengrains paved the inner wall of the locule during the unfoldingprocess, suggesting that the pollen grains bend the locule walloutward when they adhere to the wall. In the second experiment,the distribution of potassium in transverse sections of loculesin dehisced and indehisced anthers was observed. In indehiscedanthers, potassium was detected outside the pollen grains. Incontrast, in dehisced anthers, potassium was detected insidepollen grains. This suggested potassium ions moved from theinter-pollen space (locular fluid) into the pollen grains inthe locule at the time of pollen-grain swelling. Copyright 2000Annals of Botany Company Hordeum vulgare L., locule unfolding, pollen grain swelling, potassium ion, two-rowed barley     

Two-dimensional Separation of the Prolamins of Normal and High Lysine Barley (Hordeum vulgare L.)

The polypeptide components of the reduced prolamin fraction(hordein) of barley seed proteins have been separated, beforeand after alkylation, by polyacrylamide gel electrophoresisusing buffers containing urea and/or sodium dodecylsulphate(SDS). Alkylation of the protein with 4-vinylpyridine or acrylonitrileresults in a considerable sharpening of the protein bands andsome minor changes in the band pattern. The procedure has beenused to compare the hordeins of the normal commercial varieties,Julia and Bomi, to those of a high lysine mutant of Bomi (Rise,1508). Whereas the alkylated hordein fractions of Bomi and Julia containSDS bands of apparent molecular weights 13 000, 16 000, 20 000,30 000, 43 000, 51 000, 67 000, and 86 000, the mutant hordeinfractions contain predominantly the low molecular weight (13000, 16 000, and 20 000) and mol. wt. 51 000 bands. Further resolution of the fractions was obtained by two-dimensionalelectrophoresis using 6 M urea in glycine/acetate buffer atpH 4?6 as the first dimension and SDS in tris/borate bufferat pH 8?9 as the second. Separation of the Rise 1508 hordeinin this system demonstrated that the mol. wt. 51 000 band containsseveral closely similar components.