黑麦草(Lolium perenne L.)内生真菌的检测、分离及鉴定

从多年生黑麦草(LoliumperenneL.)5个品种———SR4000、Pinnacle、Topgun、CalypsoⅡ、Justus中分离出61个菌株。次培养后,所得形态稳定的菌株可分为4个形态群,依据其形态特征及APPCR的结果,确定其中的57个分离菌株为Neotyphodiumlolii。

普通小麦-黑麦草(Lolium perenne L.)易位系鉴定及其后代灌浆与产量性状分析

为明确外源染色体或片段对小麦籽粒灌浆和产量性状的影响,以普通小麦宁春4号和黑麦草远缘杂交后代为材料,借助分子细胞学手段鉴定其BC1代染色体组成,并对其BC1F3代籽粒灌浆和产量性状进行了分析。结果表明,ND99株系出现两个顶端小片段易位类型的GISH杂交信号,为真杂种;ND99的BC1F3后代中分离出有芒(ND99A)和无芒(ND99B)两种类型,并且同一个分子标记3EST-185能够追踪这两种类型。宁春4号和易位系后代(ND99A、ND99B)籽粒的体积、鲜重、干重、水分均表现为先增后降的变化趋势;宁春4号和ND99A的灌浆速率呈双峰曲线变化,而ND99B呈多峰曲线变化。宁春4号的灌浆时间早,快速增长期长,籽粒饱满指数和粒重最大;ND99A灌浆迟,持续期短,籽粒饱满指数最低,粒重仅次于宁春4号;ND99B灌浆的峰值出现较早、较大,灌浆持续期短,籽粒饱满指数居中,粒重最低。易位系后代的穗长、穗粒数和结实小穗数明显高于宁春4号,可能是影响其灌浆的关键因素之一。

Cu^2＋胁迫对黑麦草(Lolium perenne L.)去除富营养化水体中氮、磷效果的影响

为探讨不同浓度Cu2+胁迫对黑麦草去除富营养化水体中N、P营养盐效果的影响,采用小试培养的方法,研究了Cu2+存在条件下黑麦草对水体中N、P去除的动力学过程及黑麦草植株对不同浓度Cu2+胁迫的生态响应.结果表明,0.1~2.0 mg/LCu2+胁迫均降低了黑麦草对水中N、P营养盐的去除速率,其去除过程符合二阶动力学模型.黑麦草对水体中Cu2+亦有良好的去除作用,根部是积累重金属Cu的主要部位,其对Cu2+的吸收速率符合米氏动力学方程.较低浓度Cu2+(≤0.2 mg/L)胁迫对黑麦草去除N、P的效果影响不大,并且能促进黑麦草生物量的增加,而较高浓度Cu2+(≥0.5 mg/L)胁迫虽有促进根系数量增加的趋势,但明显抑制了黑麦草根伸长和叶片生长等作用,因而严重影响了黑麦草对水体中N、P的去除.

Transfer DREB into Lolium perenne L. to improve its drought tolerance

A method of Agrobacterium tumefaciens mediated transformation for perennial ryegrass was developed using the calli of ryegrass derived from mature enrbryos. The calli were inoculated with a disarmed A. tumefaciens strain EHA105 harboring binary vector p2328. Vector p2328 contained transcription factor DREB1B and neomycin phosphotransferase （npt H） genes which were driven by promoters of rd29B and CaMV35S, respectively. The inoculated calli were selected on paromomycin- or kanamycin-containing media till the established plants being transferred to soil. Six tmnsgenic plants with DREB1B had been obtained from perennial ryegrass strain Tove. PCR and Southern-blotting showed that npt Ⅱ and DREBIB genes were integrated in perennial ryegrass genome. Stress treatment confirmed that transgenic plants with higher drought tolerance were obtained.

Italian Ryegrass (Lolium perenne ssp. multiflorum) and Corn (Zea mays) Competition

Italian ryegrass is an annual/biennial grass that is typically used as a pasture crop or a cover crop along roadsides, rights-of-way, and industrial areas. Glyphosate-resistant (GR) Italian ryegrass populations have been documented around the world, mostly in orchard and vineyard situations. The first evidence of evolved GR Italian ryegrass in row/agronomic crops was reported from Washington County, Mississippi in 2005. GR Italian ryegrass populations can jeopardize preplant burndown options in reduced-tillage crop production systems, thereby, delaying planting operations. The effects of competition of Italian ryegrass on crop growth and yield are poorly understood. A field study was conducted in the 2012 growing season and repeated in the 2013 growing season. GR and susceptible (GS) Italian ryegrass populations were established in the greenhouse and transplanted in prepared corn row beds in the fall of 2011 and 2012 at 0, 1, 2, 3, and 4 plants·meter> of crop row. Italian ryegrass plants overwintered and developed over the following spring-summer. Glyphosate was applied at 1.26 kg·ae/ha (1.5× of labeled rate) in the spring to burndown the Italian ryegrass plants and corn was planted into the ryegrass residue 2 - 3 wk later. Current corn production practices were followed. Corn density (early and late season), height (early season), and yield and Italian ryegrass biomass (early-mid season) measurements were recorded during both years. Corn height was greater in 2012 than that in 2013 at comparable stages of the growing season, due to a cooler and wetter early season in 2013 than that in 2012. Averaged across weed densities, corn density (both early and late season) and yield were higher in the GS than those in the GR population, but Italian ryegrass biomass was similar for both populations. Averaged across Italian ryegrass populations, corn density (both early and late season), and yield were inversely proportional to Italian ryegrass density. In summary, Italian ryegrass significantly reduced corn density and yield and reduction was greater with the GR than that with the GS population. Studies are underway to study inter population competition in Italian ryegrass and investigate allelopathic effects of Italian ryegrass on selected crops.

Effects of Nitrogen Fertilization on Some Turfgrass Characteristics of Perennial Ryegrass (Lolium perenne L.)

The paper aimed to study the effects of nitrogen fertilization on the yield and some plant characteristics of perennial ryegrass (Lolium perenne L.). The study was conducted at Agricultural Research and Application Center of the Faculty of Agriculture in Igdir University in 2013. In the experiment, eight levels of fertilizers: 0, 10.0, 20.0, 30.0, 40.0, 50.0, 60.0 and 70.0 kg N/ha per month were applied in perennial ryegrass (Lolium perenne L.) during the sixth growing season. A randomized complete block with three replications was used as the experimental design. Nitrogen application affected plant height, green grass yield, plant covering rate, canopy color and turfgrass quality values of perennial ryegrass positively. In both monthly and as the average of nitrogen application, times of application (spring, summer and fall) caused the most uniform turfgrass quality and establishment. Results indicated that although a significant impact was on the character investigated, such as the leaf length, green grass yield, plant cover ratio, canopy color and turfgrass quality, the doses of nitrogen fertilizer have created close to each other in groups. In the stand point of view of all characters, N1, N2 and N3 were located in first group, N4 and N5 in the second group, N6 and N7 in the third group, expect for control (N0). It can be concluded that 40.0-50.0 kg N/ha per month may be more suitable to be used in ryegrass production in the landscape.

双聚材料对干旱条件下多年生黑麦草幼苗生长的影响

干旱是限制工程创面植物生长发育的关键环境因子,双聚材料(Double polymers SJ)是用于边坡土体防渗固化的新型材料,对于工程扰损区植被恢复具有重要意义。本研究选择黑麦草(Lolium perenne L.)为研究材料,采用盆栽试验,分析不同干旱条件下SJ对黑麦草生长的影响,结果表明:中、重度干旱处理显著降低了黑麦草存活率、植株含水量及叶绿素含量,增加了植株·O^(2-)、H_(2)O_(2)、MDA等毒性物质的积累。添加SJ一方面提高了土壤的保水性(蒸发量减少约50%),另一方面显著增加了黑麦草植株的含水量和叶绿素含量,并降低了活性氧等有毒物质含量,有利于黑麦草生物量的积累及存活率的提高。因此,SJ的保水性及其对植物生长的促进作用,有利于干旱区工程边坡的生态修复。

多年生黑麦草质膜型水通道蛋白基因LpAQP的克隆及功能分析

【目的】研究黑麦草在逆境胁迫下水分代谢的基因调控,克隆黑麦草质膜型水通道蛋白基因的全长cDNA序列,并对其表达模式进行研究。【方法】利用RACE技术从黑麦草(品种"顶峰")中获得LpAQP全长cDNA序列;运用生物信息学软件分析其编码蛋白;构建LpAQP与GFP融合的瞬时表达载体,采用基因枪法转入洋葱表皮细胞,观察其细胞定位;通过real time-PCR分析LpAQP的表达模式,并用Southern杂交分析其拷贝数。【结果】克隆获得LpAQP的cDNA序列,GenBank登录号为JX569791,其开放阅读框(ORF)为867 bp,编码288个氨基酸,蛋白质分子量为30.7 kD。该基因含有2个NPA单元,6个跨膜区,LpAQP含有与MIP家族完全一致的蛋白质保守区,其氨基酸序列与其它禾本科PIP类质膜型水通道蛋白同源性较高。物种间聚类分析表明LpAQP与大麦、小麦质膜型水通道蛋白同源性较高。LpAQP可能定位于细胞质膜上。通过Southern杂交分析,发现LpAQP在黑麦草基因组中以单拷贝存在。real time-PCR分析表明LpAQP在黑麦草茎部表达高于叶和根,持续干旱胁迫会导致LpAQP表达量先上调后下降。【结论】克隆获得黑麦草质膜型水通道蛋白基因LpAQP,其以单拷贝形式存在,在黑麦草根、茎、叶中均有表达,尤其是茎中表达最高。其表达受干旱胁迫影响。

NaCl胁迫下黑麦草种子萌发过程中DNA甲基化与基因表达分析

为了解黑麦草(Lolium perenne)种子在正常条件与NaCl胁迫下萌发过程中DNA甲基化动态变化以及重要盐胁迫相关基因的表达。运用甲基化敏感扩增多态性技术分析对照和150 mmol·L^(-1)NaCl处理下黑麦草种子萌发过程(0,1,2,4,7 d)的DNA甲基化水平及动态变化,并通过实时荧光定量PCR(QRT-PCR)检测8个重要的盐胁迫相关基因的表达情况。结果表明:黑麦草种子萌发过程中甲基化水平呈下降趋势,以双链甲基化方式为主,甲基化变化同时发生在编码序列和非编码序列中。NaCl处理下DNA甲基化程度高于CK,而去甲基化程度低于CK,最终导致NaCl处理下基因组净的甲基化位点数目增加。黑麦草种子耐盐萌发过程中代谢增强,8个盐胁迫相关基因表达量呈上升趋势,而NaCl胁迫延缓了种子萌发进程,在大多数时间点的基因表达低于对照。

黑麦草种子及幼苗对镉胁迫的生理响应

为了探明不同黑麦草品种对镉(Cd)富集能力的差异及其生理机制,以6个黑麦草品种为材料,研究镉胁迫对其种子萌发、生物量、镉吸收、色素含量、光合特性及抗逆特性的影响。结果表明,与正常生长条件相比,镉胁迫对不同黑麦草品种种子萌发和根芽生长具有低促高抑作用。镉胁迫增加了黑麦草光合色素含量;镉胁迫增加了维纳斯、奔驰、维多利亚叶片的气孔导度、胞间CO_(2)浓度、蒸腾速率;镉胁迫增加了奔驰、维多利亚、蓝标叶片的POD活性;镉胁迫增加了卡特、维多利亚叶片的SOD活性;镉胁迫增加了维多利亚、维纳斯叶片的CAT活性。奔驰、维多利亚、维纳斯可能较其他品种具有更强的耐镉性,从而减少镉对自身的危害;镉胁迫增加了所有品种可溶性糖含量及除卡特外的5个黑麦草品种脯氨酸含量,奔驰、蓝标、维多利亚能产生较多的渗透调节物质来减轻镉胁迫对其造成的伤害;在正常生长条件下,黑麦草地上部镉含量和镉累积量都较低,品种间差异较小。镉胁迫明显增加了黑麦草地上部镉含量和镉累积量,奔驰地上部镉含量和镉累积量最高,显著高于其他品种。综上,奔驰、维多利亚抗镉性及富镉能力较强,是镉污染土壤修复的优选品种。

镧对NaCl胁迫下黑麦草幼苗根系生理特性的影响

采用水培法研究NaCl胁迫下黑麦草幼苗根系活性氧代谢和渗透调节物质含量的变化及La(NO3)3对其变化的影响,结果表明:随着NaCl浓度的增加,黑麦草幼苗根中丙二醛(MDA)、超氧阴离子(O2.-)和过氧化氢(H2O2)含量增大,过氧化氢酶(CAT)、过氧化物酶(POD)和抗坏血酸过氧化物酶(APX)活性及抗坏血酸(ASA)、谷胱甘肽(GSH)、可溶性蛋白质和脯氨酸含量先升后降,超氧化物歧化酶(SOD)活性和可溶性糖、Na+含量不断提高,K+含量和质膜H+-ATP酶活性持续下降。添加20 mg/L La(NO3)3处理提高了NaCl胁迫下幼苗根中SOD、CAT和H+-ATP酶的活性及ASA、GSH、可溶性蛋白质、可溶性糖和K+的含量,降低了APX活性及O2.-、H2O2、MDA、脯氨酸和Na+的含量,POD活性在低浓度NaCl(50 mmol/L、100 mmol/L)时下降,高浓度(200 mmol/L、300 mmol/L)时升高。表明适宜浓度的La(NO3)3可通过提高盐胁迫下根系清除活性氧和渗透调节的能力,降低植株的膜脂过氧化,从而增强黑麦草的耐盐性。

内生菌根菌剂浓度对两种草坪草生长特性的影响

以北方地区常用的多年生黑麦草和草地早熟禾为试材,施加不同浓度的内生菌剂,浓度梯度T1为10%,T2为30%,T3为50%,T0为不施加菌剂的空白对照,以菌剂拌种的方式施加菌剂,在正常养护60 d后做侵染检测,测量不同菌剂浓度下植物的根系活力、相对电导率、可溶性蛋白质、可溶性糖和叶绿素含量的指标变化,研究了施加不同浓度的菌剂对黑麦草和草地早熟禾的影响,以期筛选出最适菌剂浓度,提高植物的生长质量。结果表明:在所试植物材料中施加菌剂浓度在50%时侵染效果最佳,草地早熟禾和多年生黑麦草侵染率可达38.71%和46.73%,其中草地早熟禾根系活力提高至13.22%,电导率降低至14%,可溶性蛋白质含量达2.91 mg·g^(-1),可溶性糖含量达28.84 mg·g^(-1),叶绿素含量可达3.10 mg·g^(-1);而多年生黑麦草根系活力提高至114.42%,电导率降低至11.00%,可溶性蛋白质含量达3.51 mg·g^(-1),可溶性糖含量达27.28 mg·g^(-1),叶绿素含量可达3.22 mg·g^(-1)。综上所述,施加50%浓度的菌剂可有效提升供试植物材料的生长质量。

毒麦及其近似种rDNA ITS-RFLP初步分析

本研究对毒麦属6个种的核糖体ITS进行PCR-RFLP分析,首次从分子水平上鉴定区分了毒麦属4个种(毒麦、亚麻毒麦、多花黑麦草、瑞士黑麦草)。根据这4个种ITS的测序结果,选取3种内切酶(BssSI、TfiI、NehI)应用于PCR-RFLP分析。根据酶切图谱可将4个近似种鉴别区分。为建立PCR- RFLP分子鉴定方法而有效地区分毒麦及近似种提供依据。

Glomus intraradices对黑麦草生长和富集镉的影响

采用盆栽方法模拟不同镉污染状况,研究接种丛枝菌根真菌(AM)Glomus intraradices对黑麦草生长和富集、运转镉的影响。结果表明,土壤镉水平增加明显提高了黑麦草菌根侵染率,而显著降低了黑麦草的生长量,但对磷浓度无显著影响,表明Glomus intraradices对镉胁迫具有一定的耐性,并对黑麦草的磷营养和生长未表现出促进作用。Glomus intraradices的存在明显促进了黑麦草对镉的富集,菌根形成强化镉在根系的固持作用,减少镉向地上部运转,进而降低黑麦草地上部的镉浓度,减轻了镉对地上部的毒害,表现在根富集系数均大于1,运转系数远远小于1。这一结果对镉污染农田修复以及草坪草和牧草品质的改善都有重要意义。

氧化石墨烯对黑麦草种子内生真菌群落结构和多样性的影响

为了探究氧化石墨烯(GO)对黑麦草种子内生真菌群落结构和多样性的影响,将黑麦草种子在0.4%、0.8%和1.2%水平的GO溶液中胁迫4 d,采用高通量测序技术,分析GO胁迫下黑麦草种子内生真菌群落组成和多样性的变化。结果显示,4个样本所有样品共分离获得303种真菌,归属于10门39纲84目160科240属。在门分类水平上,子囊菌门(Ascomycota)和担子菌门(Basidiomycota)是主要的内生真菌类群;在属分类水平上,各处理的共有优势菌属为链格孢属(Alternaria)。不同GO处理黑麦草种子内生真菌群落结构存在差异,随着GO浓度的增加,子囊菌门的丰度出现下降,0.8%和1.2%GO处理较对照分别显著降低了19%和20%(P<0.05);所有GO处理的担子菌门丰度均显著高于对照(P<0.05);1.2%处理链格孢属的丰度较对照显著降低了37.36%。与对照相比,1.2%GO处理内生真菌的丰富度和多样性显著增加,ACE、Chao1和Shannon指数分别增加了123.5%、127.4%和117.5%(P<0.05)。主坐标分析(PCoA)分析表明,1.2%GO处理内生真菌群落结构与其他处理有较大差异;线性判别分析(LEfSe)分析发现,各处理差异指示种明显不同。可见,GO改变了黑麦草种子内生真菌群落的组成和多样性,尤其是高浓度处理(1.2%)。研究可为碳纳米材料暴露对共生物种的潜在影响研究提供参考。

巨大芽孢杆菌LY02对黑麦草修复重金属污染土壤的影响

采用盆栽试验方法,研究了接种巨大芽孢杆菌LY02对黑麦草修复Cd、Cu污染土壤以及二者复合污染土壤效果的影响。结果表明:在Cd、Cu污染土壤及复合污染土壤中接种巨大芽孢杆菌LY02,可显著提高黑麦草地上部生物量,增幅达65.0%~108.3%;促进了黑麦草在3种污染土壤中对重金属的吸收,其中对Cd污染土壤中黑麦草吸收Cd的影响最为显著,地上部Cd吸收量较对照组增加了45.8%(P<0.05);3种污染土壤中黑麦草根际土壤有效磷含量显著升高,较对照组分别增加了18.2%,26.7%,16.2%;黑麦草根际土壤中有效态Fe含量显著提高,Cu单一污染土壤中增幅最大,达到152.5%;3种污染土壤中有效态重金属含量升高,在Cu污染土壤中,有效态Cu增幅达到49.7%(P<0.05)。综上所述,巨大芽孢杆菌LY02通过增加污染土壤中生物可利用态P和Fe,促进黑麦草生长;通过提高有效态Cd和Cu的含量,增加黑麦草对其吸收,从而提高了黑麦草对Cu、Cd污染土壤的修复效率。

Revegetation of Copper Mine Tailings with Ryegrass and Willow

To restore vegetation on metal mine tailings is very difficult because theyoften contain high concentrations of heavy metals, low nutrient content and low water retentioncapacity. This study involved 3 experiments that evaluated the effects of 4 treatment amendments:montmorillonite, rice straw, organic manure and chemical fertilizer on the growth of ryegrass(Lolium perenne L.) and willow (Salix viminalis L.) with Cu and Zn mine tailings from two miningareas. The results showed that ryegrass was the most tolerant of 4 crops to Cu toxicity. Also whenorganic manure, which contained high concentrations of inorganic salts, was added to the minetailings, it significantly hindered ryegrass growth (P = 0.05). Meanwhile, with ryegrass organicmanure significantly increased (P = 0.05) the extractable Cu concentration in both mine tailings.When montmorillonite was used as a mine tailings amendment with willow, the height and tress numberat the 1st cut were significantly greater (P = 0.05) than a control without montmorillonite. Howeverthere was no significant difference for height, tress number, dry weight or root dry weight at the2nd cut. So, amendment applications to reduce metal toxicity and increase nutrients retention inmine tailings were essential during revegetation of mine tailings.

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.

Genome-wide markers for seed yield and disease resistance in perennial ryegrass

The market success of perennial ryegrass(Lolium perenne L.) cultivars depends on sufficient seed production, as they are propagated by seed. However, breeding for high quality forage production reduces seed yield, and breaking the negative correlation would help to overcome the problem. The foliar disease crown rust is another factor affecting reproductive capacity and thereby seed yield. We evaluated seed yield-related traits and resistance to crown rust in a collection of commercial cultivars and ecotypes of perennial ryegrass and identified genome-wide markers associated with the traits. The study revealed high variation between the ecotype and cultivar groups as well as between years. A genome-wide association study identified 17 DNA single-nucleotide polymorphisms(SNPs) of which eight were associated with crown rust and nine with flag-leaf length. The SNP markers were located within or near predicted genes functioning in defense against pathogens. The identified genes are strong candidates for a further in-depth functional study to continue unravel determination of leaf architecture and crown rust resistance in perennial ryegrass.

哈茨木霉对黑麦草气体交换和生长发育的影响

以黑麦草为试材,采用接种和未接种哈茨木霉菌的方法,研究了哈茨木霉定殖对黑麦草的生长发育、根系浸染共生程度、叶绿素荧光参数、气体交换特性、叶绿素含量以及生物量累积的影响,以期为哈茨木霉在黑麦草生产实际中的应用提供参考依据。结果表明:1)哈茨木霉的定殖显著改善了黑麦草的气孔导度和蒸腾速率,并显著提高了水分亏缺;2)提高了黑麦草在光系统Ⅱ处可调节性的能量耗散的非光化学淬灭量子产量;3)显著增加了黑麦草的净光合速率,从而显著增加了生物量的累计,促进了分蘖。因此,哈茨木霉的定殖可以促进黑麦草的生长发育。