Effect of Chilling Stress on Synthesis of Antioxidant Enzymes, Osmotic Adjustment Substances and Membrane Lipid Peroxide Levels in Two White Clover Cultivars

Although white clover has been widely studied and applied,less is known about its physiological responses to chilling stress in northern China.The effects of acclimation time of chilling stress in the two white clover cultivars,Rivendel and Haifa,after exposure to 4℃for 0,4,8,12,24 and 48 h were investigated.Seven physiological indexes were measured in shoots of the two cultivars.It was found that Rivendel had a strong ability to maintain the stability of cells,cope with oxidative damage and accumulate osmotic adjustment substances compared with Haifa.In addition,a comprehensive evaluation on the chilling tolerance of the two cultivars based on subordinate function value analysis showed a stronger chilling tolerance of Rivendel than Haifa.

Removal of Pyrene from Contaminated Soils by White Clover

Phytoremediation has been used as an emerging technology for remediation of soil contamination with polycyclic aromatic hydrocarbons(PAHs),ubiquitous persistent environmental pollutants derived from natural and anthropogenic processes,in the last decade.In this study,a pot experiment was conducted to investigate the potential of phytoremediation of pyrene from spiked soils planted with white clover(Trifolium repens)in the greenhouse with a series of pyrene concentrations ranging from 4.22 to 365.38 mg kg-1.The results showed that growth of white clover on pyrenecontaminated soils was not affected.The removal of pyrene from the spiked soils planted with white clover was obviously higher than that from the unplanted soils.At the end of the experiment(60 d),the average removal ratio of pyrene in the spiked soils with white clover was 77%,which was 31%and 57%higher than those of the controls with or without micobes, respectively.Both roots and shoots of white clover took up pyrene from the spiked soils and pyrene uptake increased with the soil pyrene concentration.However,the plant-enhanced dissipation of soil pyrene may be the result of plant-promoted microbial degradation and direct uptake and accumulation of pyrene by white clover were only a small part of the pyrene dissipation.Bioconcentration factors of pyrene(BCFs,ratio of pyrene,on a dry weight basis,in the plant to that in the soil)tended to decrease with increase in the residual soil pyrene concentration.Therefore,removal of pyrene in the contaminated soils was feasible using white clove.

Resource Sharing by the Different Clonal White Clover Plant(Trifolium repens L.)

Resource sharing in the different parts of the clonal white clover plant (Trifolium repens L.) was investigated in this study. The results showed that the integration of water and nitrogen metabolism occured among parental, young and old branches of a plant when water passed through its stolons. Under the sufficient supply of light, water and nutrients from soil the newly developed branches showed similar capacity in taking in sufficient water and nitrogen to that of old branches. The development of the new branches did not affect the growth of the old and parental branches attached. The results also indicated that energy values in roots was significantly related with plant age, however, the energy values were various in the different parts of a plant.

White (<i>Trifolium repens</i>L.) and Arrowleaf (<i>Trifolium vesiculosum</i>Savi) Clover Emergence in Varying Loblolly Pine (<i>Pinus taeda</i>L.) Tree Alley Spacings

Agroforestry systems have the potential to provide year-long income opportunities via the integrated forage or crop, timber, and livestock. Legumes are an attractive alternative option during the growing season when more traditional forages may not be as productive. The objective of this study was to test the establishment of arrowleaf and white clover grown under varying pine tree alley widths. In 2011, existing forage was removed in 15-yr old loblolly pine tree row alleys of different widths (3.7, 4.9, 7.3, and 9.8 m), including an open area. Arrowleaf, as an annual, was replanted in 2012. Seedlings were counted twice/year, while dry matter was measured three times/year. Photosynthetically active radiation (PAR) was measured in all alley widths to compare light penetration through the canopy. Hot and dry conditions occurred throughout 2012, affecting results. In 2012 and 2013, the greatest PAR for most treatments was observed in June. Seedling counts for all treatments were greatest immediately after establishment, and gradually declined throughout the course of the study. Dry matter yields increased throughout the growing season, and were greatest in arrowleaf clover in the open area on all measurement dates;however, increased weed pressure and repeated flooding affected yields. This study demonstrated that clover establishment in shady wooded areas is possible, but only under suitable environmental conditions.

白三叶种质资源耐酸铝性综合评价

白三叶(Trifolium repens)是一种世界性优质的蛋白饲草,在我国长江流域及以南地区被广泛栽培,而南方地区多为酸性土壤。研究发现,白三叶具有一定的耐酸铝性,因此筛选耐酸铝种质对于开发白三叶优异资源、培育耐酸铝新品种具有重要意义。以30份国内外白三叶种质为对象,研究不同酸铝浓度下的白三叶萌发特性,并采用隶属函数法进行耐酸铝性综合评价。随Al3+胁迫浓度的增加,白三叶发芽率逐渐降低,发芽势和发芽指数下降,胚根变短、变粗;Al3+浓度≥1.0 mmol·L^(-1)时白三叶种质发芽率和胚根长度受到显著抑制(P<0.05)。除胚根直径外,白三叶萌发性状不同胁迫浓度间差异显著(P<0.05),且萌发期每个胁迫处理下同一性状不同白三叶材料间亦存在极显著差异(P<0.01)。在2 mmol·L^(-1)处理下白三叶多数性状具有更大的变异系数指标差异,可作为白三叶耐酸铝性综合评价的适宜胁迫浓度。聚类分析将30份白三叶划分为3个耐酸铝能力不同的类群,其种质耐酸铝性强弱与综合评价值基本一致。来自荷兰的种质‘EM196’和来自澳大利亚的种质‘EM085’具有较高的酸铝胁迫耐受性。

紫花苜蓿、白三叶硒蛋白的提取方法筛选及工艺优化

硒蛋白是微量元素硒(Se)在生物体内发挥生物学作用的主要形式,对宿主在抗氧化、抗炎、提高免疫力等方面有重要意义。为探究不同豆科牧草硒蛋白提取方法筛选及其工艺优化,本研究以紫花苜蓿(Medicago sativa)和白三叶(Trifolium repens)为原材料,采用碱提法(NaOH)、盐提法(NaCl)、酸提法(HCl)和水提法(H2O)4种溶剂提取法获得牧草硒蛋白,并在单因素试验基础上进行正交试验优化硒蛋白提取工艺。结果表明:碱提法提取紫花苜蓿和白三叶硒蛋白效果最好。紫花苜蓿最佳工艺条件为温度40℃、料液比1∶20 g·mL^(-1)、碱浓度0.15 mol·L^(-1),硒蛋白提取率为35.39%。白三叶最佳工艺条件为温度50℃、料液比1∶40 g·mL^(-1)、碱浓度0.15 mol·L^(-1),硒蛋白提取率为35.52%。本研究可为从豆科牧草中提取硒蛋白提供重要的技术支持和实践指导。

镉对不同品种白三叶(Trifolium repens L.)种子萌发及幼苗生长的影响

白三叶(Trifolium repens L.)是一种重要的多年生优质豆科牧草本,具有生物量大和耐受重金属的特点。本研究通过培养皿滤纸法,研究了不同Cd浓度(0,20,50,100μmol·L-1)对10个白三叶品种种子萌发和幼苗生长的影响,结果表明,白三叶种子的发芽势、发芽指数和活力指数随着Cd处理浓度的增加而显著降低,发芽率没有显著变化。20μmol·L-1的Cd处理显著抑制了所有参试品种的芽长,100μmol·L-1的Cd处理显著抑制了根长。利用隶属函数法对10个品种白三叶的Cd耐受性进行了综合评价,结合抗氧化系统相关生理指标(MDA,SOD,POD等)分析表明,‘Siral’以其较高的Cd耐受性,可推荐为重金属镉污染土壤种植的理想品种。

Genetic Diversity of Clover by SRAP

In the present study,genetic diversity and genetic relationship of 11materials including red clover ( Trifolium pretense L.),white clover ( T.repen L.) and alsike clover ( T.hybridum L.) and leaf-type variants of white clover were investigated by SRAP (sequence related amplified polymorphism).Forty SRAP primers were screened,and 792 reliable bands were amplified,among which 426 (53.8%) were polymorphic.The number of polymorphic bands per pair of primers ranged from 3 to 38 with an average of 10.6.SRAP cluster analysis showed that the similarity coefficients between white clover materials were from 0.465 to 0.997 with an average of 0.812,indicating that there was certain genetic diversity among white clover.Specific bands appeared in white clover variants,indicating that there were certain differences in DNA sequence between normal while clover plants and their leaf-type variants.The similarity coefficients between red clover materials were from 0.457 to 0.827 with an average of 0.597,indicating that the different red clover germplasms had genetic diversity at the DNA molecular level.The results also showed that the genetic relationship of the alsike clover with the red clover was closer than that with the white clover.This study shows that SRAP technique can be effectively used for the analysis on interspecific and intraspecific relationship,germplasm resource identification and genetic diversity of clover.

丛枝菌根真菌和根瘤菌对白三叶氮同化的影响

为揭示丛枝菌根真菌(AMF)和根瘤菌在白三叶氮(N)同化中的作用,该研究对白三叶进行单一或联合接种隐类球囊霉(Paraglomus occultum)和三叶草根瘤菌(Rhizobium trifolii),分析其对白三叶的生长、光合作用、叶片N和氨基酸含量以及N同化相关酶活性的影响。结果表明:(1)单一接种AMF或根瘤菌以及联合接种AMF和根瘤菌均显著增加了白三叶的株高、匍匐茎长度、叶片数、地上部生物量、总生物量、叶绿素b和总叶绿素含量、稳态光量子效率和叶片N含量,这种增强效应是联合接种>单一AMF>单一根瘤菌>未接种处理。(2)联合接种AMF和根瘤菌显著增加了白三叶叶片中丙氨酸、精氨酸、天冬酰胺、天冬氨酸、谷氨酰胺、谷氨酸和组氨酸的含量,显著提升了叶片N同化相关酶如硝酸还原酶、亚硝酸还原酶、谷氨酰胺合成酶、谷氨酸合成酶、谷氨酸脱氢酶、天冬酰胺合成酶和天冬氨酸转氨酶的活性,显著促进AMF对白三叶根系的侵染。综上认为,联合接种AMF和根瘤菌通过激活N同化相关酶活性有效促进N同化,产生更多氨基酸,进一步促进白三叶植株生长;联合接种AMF和根瘤菌具有协同作用,有效促进了白三叶的N同化。

蚯蚓堆肥与生物炭对瓜草复合系统土壤及植株的影响

为探究蚯蚓堆肥与生物炭对西瓜—白三叶草生境系统的作用机制,采用温室栽培试验,设置空白对照(西瓜植株,CK)、生物炭(B)、覆盖三叶草(W)、蚯蚓堆肥(V)及其组合WV、BV、WB和WBV等8个处理,比较分析蚯蚓堆肥与生物炭对西瓜—白三叶草复合系统的土壤养分、微生物群落结构及西瓜代谢组学特征的影响。结果表明,(1)与对照相比,复合处理的土壤容重降低,肥力提高,西瓜植株的养分吸收能力增强,WBV处理综合表现最优。(2)WBV处理土壤中微生物菌群丰富度相比对照下降,其中放线菌门、变形菌门、厚壁菌门细菌以及被孢霉门真菌的相对丰度明显提升,绿弯菌门、酸杆菌门和芽单胞菌门的相对丰度下降;各复合处理土壤中微生物菌群Shannon指数、Chao 1指数均表现为BV>WB>CK>WV>WBV。(3)WBV处理组西瓜果肉检测到110种差异代谢物,其中19种代谢物为WBV处理独有,包括亚麻酸、裂解型磷脂酰胆碱、磷酰肌醇酸等脂质化合物,以及丙氨酸、天冬氨酸与谷氨酸等氨基酸。研究发现,覆盖三叶草+生物炭+蚯蚓堆肥复合系统可显著提高土壤养分含量,促进西瓜对氮、磷元素吸收,同时改善土壤微生物环境,改变西瓜植株代谢水平,提升其生物膜受保护能力和细胞渗透调节能力,从而增强其抗逆能力。

胺鲜酯(DA-6)浸种对盐胁迫下白三叶种子萌发及抗盐性的影响

以白三叶(Trifolium repens L.‘Haifa’)为研究对象,研究了0.8 mmol·L-1胺鲜酯(DA-6)浸种处理对白三叶种子在盐胁迫条件下萌发的影响,并对DA-6诱导的耐盐机理进行初步研究,为白三叶的栽培管理提供参考。结果表明,DA-6浸种显著提高了盐胁迫下种子的发芽势、发芽率、活力指数、发芽指数、茎长和鲜干重(P<0.05)。盐胁迫显著诱导了游离脯氨酸和可溶性糖的积累,降低了细胞渗透势(P<0.05)。此外,盐胁迫导致了萌发种子中活性氧(超氧阴离子和过氧化氢)和膜脂过氧化产物丙二醛含量显著增加,电导率显著升高(P<0.05)。DA-6浸种前处理显著提高了胁迫下抗氧化酶(SOD、POD和APX)的活性,增幅分别为33.0%、31.9%和49.1%,这有效地缓解了由盐胁迫引起的氧化损伤,有利于幼苗稳定细胞膜(P<0.05)。本研究表明DA-6浸种处理通过增强抗氧化防御并降低细胞氧化伤害,从而提高盐胁迫下白三叶种子的萌发特性和胁迫耐受性。

基于文献计量的白三叶草栽培技术

为深入探究白三叶(Trifolium repens)在草学研究领域中的发展脉络,从主观判断转向客观计量,该研究以CNKI和Web of Science核心合集为数据来源,借助VOSviewer和CiteSpace软件对白三叶草栽培研究的文献进行计量分析。结果表明,关于白三叶栽培技术研究的发文量呈现“初期缓慢、中期迅速、后期趋减”的趋势,西北农林科技大学、四川农业大学和甘肃农业大学是国内的主要研究机构。经过生成期、转折期和成熟期3个阶段,白三叶栽培技术的研究主题逐步从牧草资源开发和生态适应性向果园生草转移,其主要研究趋势为提高牧草产量和发展草业是白三叶的研究基础,新西兰等国家在该领域发挥了重要作用;“土壤微生物多样性”、“果园生草”及“果园碳氮循环”是未来白三叶牧草栽培的研究重点。

白三叶耐铝种质筛选及耐铝评价指标分析

为评价西南地区白三叶种质耐铝性强弱,发掘优异材料,试验以采自四川、贵州地区的82份白三叶种质和生产中应用广泛的国审品种‘海法’(Haifa)、‘胡依阿’(Huia)为供试材料,使用水培方法,观测不同种质在铝胁迫(4mmol·L^(-1))15 d的生长状况,利用隶属函数法初步筛选耐铝性差异材料。试验进一步通过观测9份初筛种质铝胁迫下的生长及生理变化,采用隶属函数法计算不同种质耐铝得分,采用主成分分析法分析不同耐铝评价指标贡献率,结合根尖苏木精染色结果,对白三叶种质进行耐铝性综合评价。结果表明:铝胁迫可引起供试白三叶株高、根长、干重、绿值、相对叶片含水量、叶绿素含量、最大光能转化效率及叶片健康指数显著下降(P<0.05);相对电导率和丙二醛含量显著增加(P<0.05);根尖活性铝含量增加。材料Tr016、Tr056、Tr060、Tr062、Tr077生长指标相对值的隶属函数得分排名靠前;而Tr009、Tr021、Tr040、Tr048、Tr075得分排名靠后。综合评价结果显示,材料Tr016、Tr060具有较强的耐铝性,而Tr021耐铝性较弱;主成分分析表明,铝胁迫下白三叶相对株高、相对根长和相对干重可作为生长鉴定指标,而光合参数(叶绿素含量、最大光能转化效率、叶片健康指数)及丙二醛含量可作为生理鉴定指标。该研究可为白三叶种质资源苗期耐铝性评价和新品种选育提供参考与基础材料。

贮藏年限对白三叶种子萌发、幼苗生长及生理特性的影响

本研究探讨了白三叶(Trifolium repens L.)种子萌发、幼苗生长以及生理特性在不同贮藏年限下的变化情况,以期为白三叶种质资源保存和田间生产提供参考。试验以贮藏0~5a的白三叶种子为材料,研究其萌发特性、幼苗生长情况、相对电导率、抗氧化酶及呼吸关键酶活性的变化。结果表明:白三叶种子的萌发指标和幼苗生长指标整体上随贮藏年限的增加呈先上升后下降的趋势,均在贮藏1a时达到最大值;随贮藏年限的增加,种子相对电导率逐渐上升,抗氧化酶和呼吸关键酶活性逐渐下降。采用隶属函数法进行综合分析可知,贮藏0,1a的种子其种用价值较高。本研究结果将为白三叶种质资源的保护及实际生产应用提供理论依据。

苜蓿花叶病毒和白三叶草花叶病毒复合侵染本氏烟的发病流行因素

为明确苜蓿花叶病毒(AMV)和白三叶草花叶病毒(WCMV)在室内条件下复合侵染本氏烟(Nicotiana benthamiana)的发病流行因素,以接种磷酸缓冲盐溶液(PBS)的本氏烟为对照,通过对AMV和WCMV单独接种侵染、复合侵染的不同接种浓度、不同温度和相对湿度(RH)对本氏烟发病的影响及本氏烟中两种病毒含量变化进行了研究。结果表明最适合本氏烟发病的AMV和WCMV单独接种浓度分别为20%(AMV:PBS=1:4)和50%(WCMV:PBS=1:1),两种病毒复合侵染接种的浓度为75%和25%(AMV:WCMV=3:1);最适宜发病的温度为25℃、RH为60%,在此条件下,AMV和WCMV复合接种侵染本氏烟的平均病情指数为80.12,比单独接种AMV、WCMV的病情指数分别提高了22.36%、45.28%,其病毒含量是两种病毒单独接种本氏烟中的2.11和1.60倍。综上可见,当温度为25℃、RH为60%以及AMV和WCMV复合侵染接种浓度为75%和25%时,本氏烟发病最严重。

紫色土柑橘园长期种植白三叶后的土壤团聚体有机碳及氮素分布特征

研究柑橘园长期种植白三叶对四川丘陵区紫色土有机碳及氮素在团聚体中分布的影响,为紫色土地力提升和柑橘园肥料合理运筹提供理论依据。2014—2020年在川东北丘陵区开展5年田间定位试验,设置柑橘园清耕及柑橘园间作白三叶2个处理,2020年3月,采集0~10、10~20和20~40 cm土层土壤样品,测定水稳性团聚体及有机碳、全氮含量。结果表明,川东北丘陵区柑橘园土壤水稳定团聚体以<0.25 mm小粒级为主,2~1 mm粒级含量最低。与清耕相比,柑橘园长期种植白三叶增加了2~0.25 mm粒级团聚体含量,减少了>2、<0.25 mm粒级团聚体含量,且不同土层之间均表现出这一趋势。种植白三叶显著提高土壤0~20 cm土层及不同粒级团聚体中有机碳和全氮含量,表现为随着粒级的减小含量逐渐增加、且随土层加深呈下降趋势。除0~10 cm土层>2、2~1 mm粒级外,柑橘园种植白三叶均能提高不同土层中各粒径土壤碳氮比。综上,柑橘园长期种植白三叶增加土壤>0.25 mm水稳性大团聚体含量,补充紫色土有机碳库和氮养分含量,能够改善土壤团粒结构,提升紫色土持续生产能力。

AMV和WCMV复合侵染对本氏烟4种内源激素的影响

为明确苜蓿花叶病毒(AMV)和白三叶草花叶病毒(WCMV)混合接种协生作用以及对本氏烟中4种内源激素的影响,以接种0.02 mol/L磷酸盐缓冲溶液(PBS,pH 7.0)的本氏烟为空白对照,对AMV+WCMV(3∶1)混合接种本氏烟发病症状以及接种后不同时间本氏烟中病毒浓度和生长素(IAA)、玉米素核苷(ZR)、赤霉素(GA)、脱落酸(ABA)4种内源激素含量变化进行了研究。结果表明:AMV和WCMV混合接种后,主要表现为接种叶片褪绿、黄化、后期死亡脱落,新生叶变小、卷曲呈圆筒状、叶面呈现畸形皱缩,植株矮化;本氏烟叶中两种病毒浓度在接种后第10 d时最大,比单独接种AMV、WCMV的病毒浓度分别提高了1.70倍和1.48倍,症状加重,表明AMV和WCMV两种病毒存在复合侵染协生作用,WCMV为协生病毒,AMV为被协生病毒;AMV和WCMV混合接种后第2、6、10 d的IAA、ZR、GA含量均显著低于单独接种和对照,ABA含量则显著高于单独接种和对照,在接种后第10 d,混合接种处理的IAA、ZR、GA含量较CK分别降低了59.40%、63.39%、64.56%,而ABA含量提高了287.30%(P<0.01)。综上所述,AMV+WCMV(3∶1)混合接种使本氏烟症状表现加重,病毒浓度增加,显著降低IAA、ZR、GA含量,提高ABA含量。

苜蓿花叶病毒和白三叶草花叶病毒复合侵染对本氏烟4种抗病相关酶活性的影响

【目的】探明苜蓿花叶病毒(alfalfa mosaic virus,AMV)和白三叶草花叶病毒(white clover mosaic virus,WCMV)(3∶1)复合侵染后对本氏烟(Nicotiana benthamiana)的致病机理。【方法】试验以单独摩擦接种AMV、WCMV和PBS缓冲液(pH 7.0)的本氏烟为对照,分别采用生物化学法测定了AMV+WCMV(3∶1)复合接种的本氏烟在第2、4、6、8、10天时叶片中4种抗病相关酶的活性。【结果】当AMV+WCMV(3∶1)复合接种的本氏烟叶片中过氧化物酶(peroxidase,POD)、多酚氧化酶(polyphenol oxidase,PPO)、苯丙氨酸解氨酶(phenylalanine ammonialyase,PAL)和过氧化氢酶(catalase,CAT)活性达到最大值时,与单独接种AMV、WCMV和PBS缓冲液对照相比显著提高(P<0.01),且4种酶应激反应速度不同。【结论】AMV+WCMV(3∶1)复合侵染可加重病毒病害的发生,并显著提高寄主中与抗病性相关的POD、PAL、PPO、CAT4种酶活性。

亚热带丘陵茶园间作白三叶草的生态效应

在亚热带丘陵区1a生幼龄茶园通过连续4a的大田对比试验，研究了茶园间作白三叶草的生态效应，结果发现：茶园间作白三叶草降低了土壤容重，提高了土壤孔隙度、有机质、全N、水解N的含量、K的活性、蚯蚓的数量和生物量，消耗了部分有效P；增加了土壤关键层次（0—20cm）和关键时期（4～6月）的水分含量，延缓和缩短了夏季高温干旱与秋季持续干旱时间；获得了土壤降温时增温、保温与升温时降温的双向动态调控效果，增加了同一层次土壤温度的稳定性，延缓了高温和低温的出现时间，缩短了过度高温时间，减少了高温对茶树生长产生的影响；抑制了杂草生长，增加了天敌的种类和种群数量，减少了虫害发生率。从而形成了立地环境．茶树．伴生生物群落系统自我调节的动态平衡，促进了茶树生长，改善了茶叶品质，显著增加了茶叶产量。与清耕茶园相比，茶园间作白三叶草后，春秋茶的酚氨比分别下降了17．10％和30．90％，产量提高了32．65％。

间种白三叶草和稻草覆盖控制丘陵茶园杂草效果

开展丘陵茶园杂草生态控制,可减少对茶园土壤水分和养分的消耗,避免使用化学除草剂。连续2年的田间试验,野外应用小样方多点采样调查茶园杂草种群数量、生物量、平均株高和茶芽密度等指标,结合实验室的茶叶内含成分分析和感官评审。结果表明:茶园共有杂草16科31属31种,春季(4月)以生长速度较慢、生物量小、株高在30cm以下的杂草为主,夏季(7月)以生长快、植株高和生物量大恶性杂草为主,秋季(9月)恶性杂草开始减少,酢浆草、白花蛇舌草等生物量小、株高矮的杂草逐渐增多;稻草覆盖处理茶园春、夏和秋季3个观测时期杂草物种丰富度(S)、多样性(H)、均匀度(J)等指数均高于清耕对照,优势集中性指数(C)低于清耕对照;间种白三叶草处理春、夏和秋季3个观测时期杂草总数量和生物量均显著低于清耕对照,夏、秋季杂草平均株高显著低于清耕对照;稻草覆盖和间种白三叶草处理春季茶芽重和产量显著高于清耕对照,鲜叶中游离氨基酸含量高于清耕对照,茶叶感官评审得分高于清耕对照处理。这表明稻草覆盖和间种白三叶草能改变茶园杂草群落结构,有效控制杂草生长,增加茶叶产量和改善茶叶品质。