The Influence of Temperature on Nitrate Reductase Activity of Agrostis palustris and Cynodon dactylon

The influence of temperature was studied in relation to nitrate reductase activity of creeping bentgrass (Agrostis palustris Huds. cv. ‘Toronto’) a cool season grass and bermudagrass (Cynodon dactylon L. cv. ‘Tifgreen’) a warm season grass. Maximum nitrate reductase activity of both species occurred at 20°C. The nitrate reductase level in bentgrass leaves was reduced when grown at 35°C while bermudagrass leaves were relatively unaffected. The activity per se of the bentgrass enzyme preparation was inhibited rather than synthesis of the enzyme.     

Mechanism of Isoxaben Tolerance in Agrostis palustris var. Penncross

Previous work has demonstrated that isoxaben tolerant mutantsof Arabidopsis thaliana var. Columbia are most likely alteredat the site of isoxaben binding. The salient question becomeswhether or not species selectivity to this herbicide might alsobe a result of differential target site binding. Grasses aregenerally more tolerant to isoxaben than dicots. In this communicationwe show that Agrostis palustris var. Penncross, a grass, is83-fold more tolerant in a soil incorporation test and 170-foldmore tolerant to inhibition of glucose incorporation into cellulosethan is Arabidopsis, a dicot. Cell wall fractionation of Agrostisshows a specific effect on cellulose biosynthesis. At most,5-fold of the 170-fold tolerance exhibited by Agrostis in termsof cellulose biosynthesis can be attributed to decreased isoxabenuptake under the test conditions. Furthermore, Agrostis is unableto metabolize isoxaben to any significant degree. Therefore,we suggest that the major portion of the tolerance in Agrostismight be due to differences in isoxaben binding. Key words: Isoxaben, cellulose, Arabidopsis, Agrostis, herbicide tolerance     

Population structure and mating-type genes of Colletotrichum graminicola from Agrostis palustris

Eighty-seven isolates of Colletotrichum graminicola, mostly from Agrostis palustris, were collected in grass fields, most of which were in Ontario, Canada. Specific primers were designed to amplify the mating-type (MAT) genes and, among 35 isolates tested, all yielded a band of the expected size for MAT2. For six isolates, the MAT2 PCR products were sequenced and found to be similar to that reported for MAT2 of C. graminicola from maize. Based on 119 polymorphic bands from 10 random amplified polymorphic DNA primers, analyses of genetic distances were found to generally cluster isolates by host and geographic origin. Among 42 isolates from a grass field in Ontario, significant spatial autocorrelation was found to occur within a 20-m distance, implying that this is the effective propagule dispersal distance. Although clonal propagation was observed in the 87 isolates with 67 unique genotypes, the extent of genetic variation in local populations implies some occurrence of sexual or asexual recombination.     

Heat-Shock Response in Heat-Tolerant and Nontolerant Variants of Agrostis palustris Huds

The heat-shock response in heat-tolerant variants (SB) and non-tolerant variants (NSB) of creeping bentgrass (Agrostis palustris Huds.) was investigated. Both variants were derived from callus initiated from a single seed of the cultivar Penncross. SB and NSB synthesized heat-shock proteins (HSPs) of 97, 83, 70, 40, 25, and 18 kD. There were no major differences between SB and NSB in the time or temperature required to induce the heat-shock response. When the HSPs synthesized by SB and NSB were analyzed by two-dimensional gel electrophoresis, it was apparent that SB synthesized two to three additional members of the HSP27 family, which were smaller (25 kD) and more basic than those synthesized by NSB. Analysis of F1 progeny of NSB x SB indicated that 7 of the 20 progeny did not synthesize the additional HSP25 polypeptides. These progeny were significantly less heat tolerant than progeny that did synthesize the additional HSP25 polypeptides. The X2 test of independence (X2 = 22.45, P < 0.001) indicated that heat tolerance and the presence of the additional HSP25 polypeptides are linked traits.     

Physiological Responses of Apple Trees to Supraoptimal Root Temperature

Ungrafted apple rootstocks were grown in sand cultures at constant root temperatures between 20°C to 40°C. Temperatures of 30°C and above reduced root and shoot growth. Serious damage to the leaves occurred at 35°C and above. The O₂ consumption, CO₂ evolution and respiratory quotient (RQ) of the roots showed maximum values at 35°C. Different rootstock cultivars varied greatly in their susceptibility to damage by supraoptimal root temperatures apparently due to anaerobic respiration. The more susceptible ones differed from resistant types in the larger amount of ethanol they accumulated in their roots at supraoptimal root temperature, and the more severe reduction in the malic acid content of the roots at such temperature. Acetaldehyde was also found in roots and leaves at supraoptimal root temperatures, whereas the organic acid content of the leaves tended to decrease. Supraoptimal root temperature also caused a reduction of cytokinins in both roots and leaves accompanied by a reduction in the leaf chlorophyll content. This could be prevented by the application of kinetin or benzyladenine to the leaves. In a short experiment a rise in root temperature up to 40°C caused an increase in transpiration and a decrease in the resistance of the leaves to the passage of water vapor, whereas in prolonged experiments transpiration reached a maximum and leaf resistance a minimum at 30°C. The leaf water potential increased also with increasing root temperature. Leaf temperature increased with increasing root temperature, irrespective of increasing or decreasing transpiration rates.     

Effects of Supraoptimal Temperatures on Merulius lacrymans

The growth rate of Merulius lacrymans (Jacq.) Fr. decreases very rapidly at temperatures above optimum (about 22°C). No growth occurred above 28°C. The thermal death-time of the fungus at different temperatures above maximum was determined. At 37.5°C the thermal death-time is 4 hours. At this temperature, the respiration is not directly affected, since an appreciable respiration occurred even after 6 hours. On the basis of ¹⁴C-experiments and analysis of UV-absorbing material excreted from the mycelium at supraoptimal temperatures, it is suggested that the detrimental effects of supraoptimal temperatures on this fungus include a degradation of the nucleic acids and a subsequent leakage of the nucleotides out of the cells.     

Plant regeneration via somatic embryogenesis in creeping bentgrass (Agrostis palustris Huds.)

We have established a high-frequency plant regeneration system via somatic embryogenesis from mature seeds of creeping bentgrass (Agrostis palustris Huds). The effects of 2,4-dichlorophenoxyacetic acid (2,4-D), 3.6-dichloroo-anisic acid (dicamba) and 6-benzyladenine (BA) on callus formation and embryogenesis were evaluated. Callus produced on the Murashige and Skoog (MS) (1962) medium containing 2,4-D had low embryogenic potency. In the presence of 30 M dicamba, addition of 2.25 to 9 M BA significantly enhanced embryogenic callus formation over dicamba alone. Optimum frequency of somatic embryogenesis was achieved on MS basal medium containing 30 M dicamba and 2.25 M BA. Over 80% of somatic embryos germinated and formed plantlets on half-strength MS basal medium. These plantlets grew normally in the greenhouse.Abbreviations MS Murashige and Skoog medium - 2,4-D 2,4-dichlorophenoxyacetic acid - BA 6-benzyladenine - dicamba 3, 6-dichloro-o-anisic acid     

Growth of Agrostis palustris in subsurface black-layered sand induced by cyanobacteria and sulfate-reducing bacteria

The growth of Agrostis palustris was evaluated in sand columns in response to black-layer formed by the interaction of cyanobacteria in the genera Oscillatoria (isolates OS-1 and OS-2) and Nostoc (NS-1) with the sulfate-reducing bacterium Desulfovibrio desulfuricans. All plants of A. palustris transplanted into black-layered sand columns survived, and the black-layer cleared adjacent to roots as they grew down in the column. Black-layer remained in the columns below the advancing root tips. After 10 weeks of growth, numerous roots showed discontinuous reddish-brown discoloration on their surfaces. Shoot growth of A. palustris was reduced in response to all cyanobacteria and D. desulfuricans isolates alone or in combination. Root growth was unaffected by the microorganisms with the exception of stimulation by OS-1 and inhibition by D. desulfuricans. Interaction of the microbes and the formation of black-layer is discussed relative to the growth of A. palustris. Journal Paper J-14483 of the Iowa Agriculture and Home Economics Experiment Station, Ames, IA 50011. Project 2616 Journal Paper J-14483 of the Iowa Agriculture and Home Economics Experiment Station, Ames, IA 50011. Project 2616     

剪股颖愈伤组织诱导与植株再生

以匍匐剪股颖的成熟种子为外植体,对其愈伤组织诱导及再生体系进行了研究。结果表明:愈伤组织诱导合适的培养基为MS 6mg.L-1 2,4-D 0.2mg.L-1 TDZ 500mg.L-1 CH,诱导率达到68.1%;MS 5mg.L-1 2,4-D 0.1 mg.L-1TDZ 500mg.L-1 CH为愈伤组织继代较合适的培养基;愈伤组织分化的合适培养基为MS 0.3mg.L-1 TDZ 0.5mg.L-1 6-BA,分化率达到52.7%。随着愈伤组织继代次数的增加,胚性愈伤组织的分化能力没有明显的降低,这可为后续的遗传转化长期提供受体材料。     

Transgenic plants of turfgrass (Agrostis palustris Huds.) from microprojectile bombardment of embryogenic callus

Transgenic creeping bentgrass (Agrostis palustris Huds., cv. Pencross; Poaceae) plants have been obtained by microprojectile bombardment of and regeneration from embryogenic calli with a vector designed to deliver the -glucuronidase (GUS) gene under the control of rice actin 1 5' regulatory sequences. Southern analysis of polymerase chain reaction (PCR)-amplified and restriction-digested genomic DNA of four transgenic plants regenerated from these cultures showed the unscrambled integration of the gus fragment. Northern blot analysis confirmed the expression of gus mRNA in one of the transgenic plants. Western blot analysis revealed a high level of accumulation of gus protein. Histochemical assays showed enzymatic activity of -glucuronidase in all parts of the transgenic turfgrass plant. The order of gus expression level in different tissues of the transgenic plant is as follows: stem node > first young leaf > root tip > second / third / fourth young leaf > stem internode > root hair-zone.Abbreviation GUS -glucuronidase - MS Murashige and Skoog(1962) medium - BA 6-benzyladenine - dicamba 3, 6 -dichloro-o-anisic acid - PCR polymerase chain reaction     

Nutrient salts and the toxicity of black-layer induced by cyanobacteria and Desulfovibrio desulfuricans to Agrostis palustris

Cyanobacteria and Desulfovibrio desulfuricans can interact to form a subsurface black-layer in high-sand content golf greens that impairs internal water drainage and results in the decline of the Agrostis palustris turfgrass on the green. Research was initiated to evaluate the effect of mineral salts (sulfur, iron, lime) and fructose (a soluble carbohydrate) added to a balanced nutrient salts control solution on the development and toxicity of black-layer to the growth of A. palustris. The various nutrient salts combinations were applied to single isolates of cyanobacteria and D. desulfuricans in nonblack-layered sand, and to the combination of cyanobacteria and D. desulfuricans (necessary for black-layer development) in black- layered sand. Dry weights of A. palustris treated with the salts control decreased with individual isolates of cyanobacteria and more so in the blackened sand produced by the combinations of cyanobacteria and D. desulfuricans. The addition of sulfur to the salts control increased dry weights of A. palustris growing with single isolates of cyanobacteria and in the sand blackened by the combinations of cyanobacteria and D. desulfuricans compared with the salts control; dry weight decreased in response sulfur only in nonblackened sand with D. desulfuricans alone. The addition of iron to the salts control produced the greatest increase in dry weight relative to the salts control among all single and combined microorganisms, except for D. desulfuricans. The addition of lime or fructose to the salts control decreased dry weight among plants growing in the no-organism control, with D. desulfovibrio alone, and with individual isolates of cyanobacteria relative to the salts control. Dry weights in response to lime and fructose in sand blackened by the combination of cyanobacteria and D. desulfuricans remained unchanged or decreased relative to the salts control. Growing roots of A. palustris cleared the blackening in sand and showed gold-colored cortical cells with blackened cell walls and vascular cylinders. The observations are discussed relative to the role of the various salts on the toxicity of D. desulfuricans to A. palustris in black-layered and nonblack-layer sand.     

Definition of leaf health in Agrostis palustris at the time of infection and colonisation by Curvularia lunata

The state of leaf health of intact Penneagle creeping bentgrass leaves into which Curvularia lunata was able to ingress was determined by reducing cuticle/wax formation with trichloroacetic acid (TCA) and stressing plants with high air temperatures. Leaf health was estimated throughout the growing period of the plants by extracting chlorophyll a and then regressing the values with respect to time. In this manner, leaves at each nodal position could be classified as either juvenile, mature or senescent. The addition of TCA to plants decreased leaf and plant life. Stressing plants with high air temperatures accelerated leaf senescence as did higher levels of TCA. Inoculation with Drechslera sorokiniana resulted in lesion formation within 2 days. Symptoms commonly attributed to Curvularia blight were present on plants treated with 140 ^M TCA when high-air-temperature stressed and inoculated with C. lunata. Histochemical procedures failed to show the presence of mycelium of C. lunata within the cells of Curvularia blight symptom areas. Results demonstrated that C. lunata had the ability to colonise heat stressed and/or old leaves but did not have the ability to infect and colonise juvenile or mature tissues.     

Plant regeneration from protoplasts isolated from embryogenic suspension cultures of creeping bentgrass (Agrostis palustris Huds.)

A rapidly growing embryogenic suspension culture cell line of creeping bentgrass cv Penncross (Agrostis palustris Huds.) was established from callus derived from the culture of mature seeds. High concentrations of 2,4-D were required for the induction of callus (3 mg/1) as well as for the maintenance of the cell Une (2 mg/1) on modified B5 medium of Gamborg. Protoplasts isolated from the suspension cultures were successfully cultured in Murashige and Skoog's basal medium supplemented with only 0.1 mg/1 2,4-D. Although protoplast plating efficiency was rather low (0.36%), 30% of the protocalli formed normal green plants that were successfully established in soil.Abbreviations BA 6, benzylaminopurine - 2,4-D 2,4 dichlorophenoxyacetic acid - MES 2, (N-morpholino)-ethane sulfonic acid - MS Murashige and Skoog medium (1962) - B5 Gamborg medium (1968)     

Promoter analysis in transient assays using a GUS reporter gene construct in creeping bentgrass (Agrostis palustris)

Transient expression profiles for several chimeric beta-glucuronidase (GUS) gene constructs were determined in tissues (young leaves, mature leaves and roots) of creeping bentgrass (Agrostis palustris, cv. Penn A4) following microprojectile bombardment. The constructs analyzed consisted of the uidA (GUS) reporter gene driven by four different promoters (ubiquitin 3-potato, ubiquitin corn, ubiquitin rice and CaMV 35S). The total number of GUS hits (or transient expression units; TEUs) were determined manually under a dissecting scope after histochemical staining for GUS. Results suggest that the ubiquitin rice promoter is most active in cells of turfgrass, regardless of the developmental stage or tissue-type. The ubiquitin corn promoter was the next best. Of the four promoter used, except for ubiquitin 3-potato, reporter gene activity was dramatically higher in mature leaves compared to young leaves. The relative efficiency of each promoter was about the same in roots and leaves. We have also analyzed uidA (GUS) reporter gene activity following microprojectile bombardment in transient expression assays with callus from two cultivars (Providence or Penn A4) of creeping bentgrass. Differences in the frequency of GUS positive hits were observed between cultivars up to 72 hours post-bombardment. However, this difference between cultivars disappeared after 72 hours post-bombardment. This information describing promoter functionality in bentgrass will be important when designing gene constructs for trait modification and when choosing appropriate cultivars for improvement through gene transfer experiments. This is the first in depth report on organ-specific and developmental gene expression profiles for transgenes in a turfgrass species.     

Cell-Dependent Differences in the Production of Infectious Herpes Simplex Virus at a Supraoptimal Temperature

The replication of herpes simplex virus (HSV) was compared in rabbit and hamster cells at optimal and supraoptimal temperatures. Replication occurred in cells of either species at 33 C, but the total infectious virus yield was routinely about 10-fold greater in rabbit cells than in hamster cells. At 39 C, this difference was exaggerated to greater than 100,000-fold. Whereas infectious virus was produced and plaques formed in rabbit kidney cell monolayers at the higher temperature, neither developed in those derived from hamster embryos. Elevating the temperature from 33 C to 39 C at various time intervals after exposure of the cultures to virus revealed that production of infectious virus in hamster cells was completely heat-sensitive up to 6 hr after infection. Specific viral antigens and viral deoxyribonucleic acid (DNA) were synthesized in both rabbit and hamster cell cultures. In addition, cellular DNA synthesis was depressed and cytopathic effects occurred in both cell systems. These cytopathic effects were not observed in cell cultures treated with HSV previously inactivated with ultraviolet light. Compared with parallel cultures at 33 C, the amount of viral DNA synthesized at 39 C was greatly reduced in both systems. In hamster cells, the reduction was twofold greater than in rabbit cells. This cell-dependent thermal inhibition of HSV replication in hamster cells did not occur with vaccinia virus.     

Arsenate tolerance in Agrostis castellana and Agrostis delicatula

Experiments were made to elucidate why many field-layer plants in beech forests do not grow in mor (raw humus), a layer of organic matter in various stages of decay containing little or no mineral soil particles. Three possibilities were considered directly or indirectly: (i) tree root competition and litter shading, (ii) phytotoxic and nutrient complexing organic compounds in the soil solution, and (iii) high H-ion concentration in the solution. Rhizomes of eight species (Allium ursinum, Carex sylvatica, Convallaria majalis, Deschampsia flexuosa, Galium odoratum, Poa nemoralis, Stellaria holostea, Stellaria nemorum) were collected in late winter and used in a soil experiment to test reestablishment and growth in untreated mor (soil solution pH 3.6) and after pH was raised to 4.3 by addition of SrCO₃. A flowing solution experiment was used with five of the species mentioned. The solution was composed according to the soil solution of the untreated mor but lacking organic compounds. pH was adjusted to and maintained at 3.6 and 4.3. The experiments showed that even when root competition, litter shading and organic compounds were excluded as limiting factors, only Deschampsia flexuosa, and partly Convallaria majalis, could produce new roots in the untreated mor or in the pH 3.6 solution treatment. Some shoot biomass developed in all species except in Galium odoratum, though significantly less than at pH 4.3.It was concluded that high H-ion concentration of the soil solution precludes establishment and growth of many forest plants in beech forests developed on mor podzols.