Physiological and Biochemical Responses of Perennial Ryegrass Mixed Planting with Legumes under Heavy Metal Pollution

In artificially controlled pot experiments,perennial ryegrass was mixed with other leguminous plants(white clo-ver and alfalfa)and treated with lead,zinc and cadmium(337 mg·kg^(-1),648 mg·kg^(-1),and 9 mg·kg^(-1),respectively)to simulate compound pollution conditions.The results showed that the concentrations of heavy metals,trans-port factors,and bioconcentration factors in mixed planting of ryegrass decreased compared with those in mono-culture.Regardless of whether heavy metal pollution was introduced,mixed planting increased the aboveground and underground biomasses of ryegrass.The different mixed planting treatments had no significant impact on the chlorophyll concentration of ryegrass.The mowing time,mixed planting treatment,and heavy metal treatment had impacts on antioxidant and osmotic adjustment substances,and there were some interactions.The mixed planting treatment did not significantly affect glutathione concentration,cysteine concentration,or nonprotein thiol.Mixed planting generally increased the nitrogen and phosphorus concentrations of ryegrass while reducing the stoichiometric ratio of carbon,nitrogen,and phosphorus.These results suggest that the mixed planting of ryegrass with legumes promotes the growth of ryegrass in the presence of high concentrations of heavy metal pollution.However,it does not enhance the ability of ryegrass to remediate heavy metal pollution in the soil.

Callus Induction and Plant Regeneration from Mature Seeds of Perennial Ryegrass

Objective The aim was to explore callus induction and plant regeneration of perennial ryegrass, as well as provide the foundation for transgenic research on perennial ryegrass.[ Methed] Mature seeds of perennial ryegrass were used as explants to study the effects of different hormone compositions on callus induction, proliferation and plant differentiation. Result The result showed that the induction rate achieved its highest on 2,4-D of 8 mg/L combining with 6-BA of 0.025 mg/L, which was up to 56.42%. Callus were differentiated after two to three generations, the highest differentiation rate 34.14% was achieved in the medium contained MS medium with 6-BA of 2 mg/L, and the differentiation rate was obviously affected by the callus condition after proliferation. The root inducing medium, containing 0.5 mg/L NAA and MS medium with half of macroelement, gained 98% root inducing rate. Conclusien A high frequency genetic regeneration system was established.

Scarabaeid Larvae- and Herbicide-Resistant Transgenic Perennial Ryegrass (Lolium perenne L.) Obtained by Agrobacterium tumefaciens-Mediated Transformation of cry8Ca2, cry8Ga and bar Genes

Insect pest and weeds are two major problems for forage and turf grasses. In this study, scarab larvae- and herbicide-resistant transgenic perennial ryegrass （Lolium perenne L.） was obtained by transforming it with cry and bar genes simultaneously via the Agrobacterium-mediated method. To optimize the callus induction and plant regeneration conditions, various concentrations of 2,4-dichlorophenoxyacetic acid and 6-benzylaminopurine were assayed. The transformation efficiencies of different Agrobacterium suspension media, used during Agrobacterium-mediated transformation, were compared. Then, plasmids of pCAMBIA3301 containing cry gene （cry8Ca2 or cry8Ga） and bar gene, driven by ubiquitin promoter, were transformed into perennial ryegrass. The transformants were generated and confirmed by both Southern hybridization analysis and Western hybridization analysis. Further, the resistance of transgenic perennial ryegrass plants to scarab larvae and herbicide were analyzed. After 30 d of co-cultivation with scarab larvae, the damage to the root system of transgenic plants was less than that of non-transgenic control plants. Additionally, the leaves of transgenic plants were resistant to Basta, while leaves of the wild plants wilted after Basta spraying. These results show that cry gene and bar gene were successfully transferred into perennial ryegrass by the Agrobactgerium-mediated method, and convey resistance to scarab larvae and herbicide in transgenic perennial ryegrass plants.

Selection of Rhizosphere Phosphate-solubilizing Bacteria of Perennial Ryegrass

The research isolated phosphorus-soluble bacteria from different parts of ryegrasses and selected 7 bacteria performing better in solbilizing capacity. The test results showed that the capacity of phosphorus-solubilizing tended to be volatile in the range from 135.27 to 187.87 μg/ml and the secreting capacity of IAA was in3.47-24.27 μg/ml. It is believed that Lp59, Lp61, Lp65, Lp69, Lp70 and Lp72 are potential for further development.

The LpHsfA2-molecular module confers thermotolerance via fine tuning of its transcription in perennial ryegrass(Lolium perenne L.)

Temperature sensitivity and tolerance play a key role in plant survival and production.Perennial ryegrass(Lolium perenne L.),widely cultivated in cool-season for forage supply and turfgrass,is extremely susceptible to high temperatures,therefore serving as an excellent grass for dissecting the genomic and genetic basis of high-temperature adaptation.In this study,expression analysis revealed that LpHsfA2,an important gene associated with high-temperature tolerance in perennial ryegrass,is rapidly and substantially induced under heat stress.Additionally,heat-tolerant varieties consistently display elevated expression levels of LpHsfA2 compared with heat-sensitive ones.Comparative haplotype analysis of the LpHsfA2 promoter indicated an uneven distribution of two haplotypes(HsfA2^(Hap1) and HsfA2^(Hap2)) across varieties with differing heat tolerance.Specifically,the HsfA2^(Hap1) allele is predominantly present in heat-tolerant varieties,while the HsfA2^(Hap2) allele exhibits the opposite pattern.Overexpression of LpHsfA2 confers enhanced thermotolerance,whereas silencing of LpHsfA2 compromises heat tolerance.Furthermore,LpHsfA2 orchestrates its protective effects by directly binding to the promoters of LpHSP18.2 and LpAPX1 to activate their expression,preventing the non-specific misfolding of intracellular protein and the accumulation of reactive oxygen species in cells.Additionally,LpHsf A4 and LpHsf A5 were shown to engage directly with the promoter of LpHsfA2,upregulating its expression as well as the expression of LpHSP18.2 and LpAPX1,thus contributing to enhanced heat tolerance.Markedly,LpHsfA2 possesses autoregulatory ability by directly binding to its own promoter to modulate the self-transcription.Based on these findings,we propose a model for modulating the thermotolerance of perennial ryegrass by precisely regulating the expression of LpHsfA2.Collectively,these findings provide a scientific basis for the development of thermotolerant perennial ryegrass cultivars.

Removal of nitrogen from wastewater with perennial ryegrass/artificial aquatic mats biofilm combined system

To develop a cost-effective combined phytoremediation and biological process, a combined perennial ryegrass/artificial aquatic mat biofilm reactor was used to treat synthetic wastewater. Influent ammonium loading, reflux ratio, hydraulic retention time （HRT） and temperature all had significant effects on the treatment efficiency. The results indicated that the effluent concentration of ammonium increased with increasing influent ammonium loading. The reactor temperature played an important role in the nitrification process. The ammonium removal efficiency significantly decreased from 80% to 30%-50% when the reactor temperatttre dropped to below 10℃. In addition, the optimal nitrogen removal condition was a reflux ratio of 2. The nitrate and ammonium concentration of the effluent were consistent with the HRT of the combined system. The chemical oxygen demand （COD） removal efficiency was at a high level during the whole experiment, being almost 80% after the start-up, and then mostly above 90%. The direct uptake of N by the perennial ryegrass accounted for 18.17% of the total N removal by the whole system. The perennial ryegrass absorption was a significant contributor to nitrogen removal in the combined system. The result＇illustrated that the combined perennial ryegrass/artificial aquatic mat biofilm reactor demonstrated good performance in ammonium, total N and COD removal.

Determining the earliest growth stage to detect the presence of endophytes in tall fescue and perennial ryegrass seedlings using molecular markers

Background:Tall fescue(Festuca arundinacea[Schreb.],Lolium arundinaceum[Schreb.]Darbysh)and perennial ryegrass(Lolium perenne)are important cool-season forage and amenity grasses that have a mutualistic association with an endophytic fungus.Endophytes confer insect and drought resistance to plants but can produce mammalian toxins.Novel endophytes that do not produce mammalian toxins have been introduced to elite cultivars for commercial production.Seed companies need to maintain adequate levels of novel endophytes within the elite forage cultivars.Endophyte detection is performed using immunochemical and molecular techniques because of their speed and reliability.Early detection in seedlings is essential to evaluate the viability of the endophyte within seed lots.Methods:This research aimed to identify the earliest growth stage in which immunochemical and molecular methods can detect viable endophyte in seedlings of tall fescue cultivars BarOptima(e34),Texoma MaxQII(584),and Jesup MaxQ(542),as well as the perennial ryegrass cultivar Remington(NEA2).Results:Immunochemical testing detected endophytes in seedlings 14 days after germination(DAG),but the detection rate increased until 42 DAG in some cultivars tested.The molecular marker Tef1exon detected endophytes at a lower rate than the immunochemical method at 28–42 DAG.However,there was insufficient DNA to detect endophytes in 14 DAG seedlings using markers.Conclusions:We conclude that the most accurate detection of viable endophytes in seedlings was 42 DAG,at which sufficient and consistent endophyte colonization occurred.

Genomic prediction of seasonal forage yield in perennial ryegrass

Background:Genomic selection has the potential to accelerate genetic gain in perennial ryegrass breeding,provided complex traits such as forage yield can be predicted with sufficient accuracy.Methods:In this study,we compared modelling approaches and feature selection strategies to evaluate the accuracy of genomic prediction models for seasonal forage yield production.Results:Overall,model selection had limited impact on predictive ability when using the full data set.For a baseline genomic best linear unbiased prediction model,the highest mean predictive accuracy was obtained for spring grazing(0.78),summer grazing(0.62)and second cut silage(0.56).In terms of feature selection strategies,using uncorrelated single-nucleotide polymorphisms(SNPs)had no impact on predictive ability,allowing for a potential decrease of the data set dimensions.With a genome-wide association study,we found a significant SNP marker for spring grazing,located in the genic region annotated as coding for an enzyme responsible for fucosylation of xyloglucans—major components of the plant cell wall.We also presented an approach to increase interpretability of genomic prediction models with the use of Gene Ontology enrichment analysis.Conclusions:Approaches for feature selection will be relevant in development of low-cost genotyping platforms in support of routine and cost-effective implementation of genomic selection.

Dissipation of polycyclic aromatic hydrocarbons and microbial activity in a field soil planted with perennial ryegrass

Dissipation and plant uptake of polycyclic aromatic hydrocarbons （PAHs） in contaminated agricul- tural soil planted with perennial ryegrass were investigated in a field experiment. After two seasons of grass cultivation the mean concentration of 12 PAHs in soil decreased by 23.4% compared with the initial soil. The 3-, 4-, 5-, and 6- ring PAHs were dissipated by 30.9%, 25.5%, 21.2%, and 16.3% from the soil, respectively. Ryegrass shoots accumulated about 280 ug.kg1, shoot dry matter biomass reached 2.48 x 104kg-ha1, and plant uptake accounted for about 0.99% of the decrease in PAHs in the soil. Significantly higher soil enzyme activities and microbial community functional diversity were observed in planted soil than that in the unplanted control. The results suggest that planting ryegrass may promote the dissipation of PAHs in long-term contaminated agricultural soil, and plant-promoted microbial degradation may be a main mechanism of phytoremediation.

Effects of Atmosphere Deposition on Nitrogen Content of Two Poaceae Plants

[Objective] It was to study nitrogen use efficiency under the condition of deposition of perennial ryegrass (Lolium perenne L.) and barley (Hordeum vulgare L.), further revealing the difference in gene variation.[Method] A pot experiment was conducted under 3 treatments of rainwater, ammonium-N (aN) and nitrate-N (nN).[Result] In the treatments of aN and nN, the biomass and N contents in plants were obviously higher than those in rainwater treatment, while the largest biomass(36.116 g) was observed in nN treatment. The absorption to aN was best to perennial ryegrass while nN to barley. According to the differential analysis of N content, treatments of both aN and nN was nitrogen superfluous, which led to release excessive N to the atmosphere, and the rainwater treatment and the control were N deficient, the largest N absorption from atmosphere was 0.698 g in rainwater treatment.[Conclusion] Average N use efficiency contributed by atmosphere N deposition ranged from 1.321%-6.116%, while the control of barley had the highest of 6.116%.

Some Evidence for Host Specificity in Arbuscnlar Myc-orrhizas

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Forage Species Suitability Mapping for China Using Topographic, Climatic and Soils Spatial Data and Quantitative Plant Tolerances

Selecting plants adapted to the climatic and soil conditions of specific locations is essential for environmental protection and economic sustainability of agricultural and pastoral systems. This is particularly true for countries like China with a diversity of climates and soils and intended uses. Currently, proper species selection is difficult due to the absence of computer-based selection tools. Climate and soil GIS layers, matched with a matrix of plant characteristics through rules describing species tolerances would greatly improve the selection process. Better matching will reduce environmental hazards and economic risks associated with sub-optimal plant selection and performance. GIS-based climate and soil maps have been developed for China. A matrix of quantitative species tolerances has been developed for example forage species and used in combination with an internet map server that allows customized map creation. A web-based decision support system has been developed to provide current information and links to original data sources, supplementary materials, and selection strategies.

Effects of daughter tiller removal on shoot and root growth of the parent tiller in Lolium perenne

Background:This study investigated the effects of daughter tiller removal on parent tiller development in Lolium perenne.Methods:Plants of L.perenne were grown hydroponically to allow separation of roots by phytomer position on the tiller axis and allowed to form two daughter tillers.In Experiment 1,adult daughter tillers were excised and effects on subsequent main tiller growth were observed for 16 days,on average.In Experiment 2,the growth of main tillers with or without daughter tillers was compared over 90–100 days.Two cultivars,‘Alto’bred from New Zealand germplasm and‘Aberdart’bred from United Kingdom germplasm were tested.Results:Excision of adult daughter tillers reduced the dry weight of older roots at the base of parent tiller axes and accelerated new phytomer appearance.Preventing tillers from forming daughter tillers by new tiller excision resulted in increased individual leaf and root dry weight in tillers without daughter tillers.Conclusions:The data indicated that daughter tillers contributed substrate for root development of their parent tillers.The presence of daughter tillers reduced the size of their main tiller but greatly enhanced whole plant yield.