转录组解析白三叶根际溶磷菌株RW8的解磷机制

采用转录组测序的方法分析了白三叶根际溶磷菌RW8在含有难溶磷(A组)、可溶磷(B组)与无磷(C组)培养条件下差异基因表达。以可溶磷为对照,在难溶磷条件下,分别检测到4782个基因在RW8中上调表达,447个基因下调表达;在无磷组中,共检测到3630个基因上调表达,209个基因下调表达。GO基因注释发现RW8在A组和C组中的差异基因聚类基本相同。生物学过程主要聚类在代谢过程、细胞过程、单细胞过程、刺激应答、定位以及生物反应调节;细胞组分主要聚类在细胞组分、细胞膜、膜组分与高分子配合体等;分子功能主要聚类在催化活性、结合功能与转运功能。代谢途径分析发现2-α-氧代羧、α-亚麻酸、五碳二元酸、脂肪酸、甘氨酸-丝氨酸-蛋氨酸以及缬氨酸-亮氨酸-异亮氨酸等代谢途径的基因显著被富集。挑选10个差异基因并用荧光定量检测其在A、B、C 3种不同培养条件下的基因表达,发现所有挑选基因的表达变化与转录组结果变化趋势相同。液相色谱检测有机酸组成及含量,发现在A组和C组中乳酸、琥珀酸与柠檬酸显著高于B组,富马酸与苹果酸的含量在C组中显著升高, A组与B组差异不显著;α-酮戊二酸的含量在A组中显著增高, B组和C组差异不显著。     

引进红三叶种质资源遗传多样性的ISSR分析

为了分析从俄罗斯、美国和加拿大引进的31份红三叶(Trifolium pratense)种质资源的遗传多样性,为其进一步利用提供参考,本文采用ISSR分子标记技术对其进行了遗传多样性研究。从20条ISSR引物中筛选出多态性明显、重复性好的5条引物进行扩增,共扩增出61条谱带,其中多态性条带58条,多态性比率(PPB)高达95.08%。POPGENE分析结果表明,红三叶种质间遗传相似系数(GS)变化范围在0.5902~0.9344之间,平均Shannon信息指数(I)为0.3427,平均Nei’s基因多样性指数(H)为0.2092,平均有效等位基因数(Ne)1.3162,表现出丰富的遗传多样性。利用UPGMA聚类分析,可将31份红三叶种质分为5大类,其中来自加拿大的24号种质材料与其他材料间遗传分化最大,单独聚为一类。本文还比较了ISSR标记的聚类结果与基于形态特征的聚类结果之间的异同。     

三种化学物质诱导白三叶草叶斑病抗性的研究

为了探明硅酸钾、硅酸钠、水杨酸对白三叶草(Trifolium repens)抗叶斑病的诱导效果,采用离体、活体试验及生物化学方法测定了3种化学物质对白三叶草叶斑病的室内抑菌活性、诱导抗病效果、防御酶活性及田间诱导防病效果。结果表明:浓度为25μg·mL~(-1)时3种化学物质的抑菌率较低,最高仅为4.59%,但诱导抗病效果较高,均在45.00%以上;叶片中多酚氧化酶(PPO),过氧化物酶(POD),苯丙氨酸解氨酶(PAL)活性均有明显增加,除硅酸钾、硅酸钠诱导处理的PPO在第3d达到峰值外,其他化学物质诱导处理的酶活性均在第5d达到最大值;田间处理7d后3种化学物质诱导防病效果均达到了72.00%以上;处理14d后有所降低,最低为23.08%。综上所述,浓度为25μg·mL~(-1)的3种化学物质能够激发白三叶草叶片中与抗病性相关的PPO,POD和PAL活性,对白三叶草抗叶斑病均有较好的诱导防病作用。     

无机磷溶解菌RW8的筛选、鉴定及对白三叶促生效果研究

从白三叶根际分离筛选9株溶磷微生物,并对挑选菌株进行形态、细胞以及种属鉴定,并分析其对白三叶的促生效果。通过分析溶磷圈与溶磷菌落直径的大小与比值获得溶磷量与持续溶磷能力较强的RW8菌株。通过形态与扫描电镜观察发现RW8是具有鞭毛的短杆菌,其菌落形态表面光滑,不透明;将RW8 16S rDNA序列比对NCBI数据库发现其与阴沟肠杆菌属(Enterobacter sp.)的同源性高达99.5%,Biolog鉴定其与阴沟肠杆菌(Enterobacter cloacae ss dissolvens)的相似性为0.610。RW8溶磷量达424.85 μg/mL,溶磷能力可能与RW8的产酸性能(12.30 μg/mL)密切相关。RW8对白三叶根系与幼苗的生物产量的分配有显著影响,接种RW8后能抑制白三叶根系伸长,根系鲜重与干重有下降趋势,但差异不显著;而幼苗的生物产量(干重与鲜重)均显著高于对照(P<0.05),并且该现象与植物生长激素IAA无关,其具体的作用机制还有待于进一步研究。     

秋水仙素诱导草木樨的研究

[目的]鉴定秋水仙素诱导后的草木樨的倍性。[方法]以草木樨萌动种子为试材,秋水仙素溶液为处理试剂,进行了秋水仙素诱导草木樨的研究。[结果]用浓度为0.2%的秋水仙素溶液处理草木樨能使草木樨诱导加倍,草木樨植株表现出多倍体具有的相应特征,整体显示出巨大性。[结论]草木樨为加倍成功的多倍体。     

添加剂对白三叶青贮的影响

为探讨不同添加剂对白三叶（Trifolium repens）青贮的影响,以营养生长期白三叶为原料,分别设计对照、添加甲酸（6 mL/kg）、蔗糖（2%）处理,袋装密封青贮360 d后取样分析。结果表明,添加甲酸或蔗糖都能显著改善白三叶青贮饲料的发酵品质,极显著降低青贮饲料的pH值、乙酸和氨态氮含量（P<0.01）,极显著增加乳酸含量（P<0.01）。添加甲酸或蔗糖显著提高白三叶青贮饲料的干物质保存率（P<0.05）,显著提高ADF含量（P<0.05）。青贮之后,硝酸盐含量下降。     

Host resistances to Aphanomyces trifolii root rot of subterranean clover: first opportunity to successfully manage this severe pasture disease

Subterranean clover (Trifolium subterraneum) is an important pasture legume in Australia (29 million ha) and elsewhere. However, severe pasture decline occurs in association with several root pathogens, including Aphanomyces trifolii, that has been misidentified for decades as A. euteiches until recently confirmed as A. trifolii. A series of controlled environment experiments was undertaken to identify host resistance to A. trifolii in subterranean clover and to compare virulence and phylogeny of isolates. In experiment 1, Dalkeith, Bacchus Marsh, Riverina and Yarloop were the most resistant of 38 cultivars with a percentage disease index (PDI) ≤10 for both tap and lateral roots. Experiment 2 confirmed resistance of Yarloop, but a change in some relative varietal resistances suggested physiological specialization among A. trifolii isolates. Experiment 3 confirmed extensive variation in virulence and physiological specialization across 23 isolates of A. trifolii, with three distinct clades, two of which were distinct from isolates collected previously. Experiment 4 identified host resistance(s) effective against a mixture of 20 A. trifolii isolates, but the most resistant cultivars (Antas, Uniwager, Leura) still showed significant disease. This is the first study to show physiological specialization in A. trifolii and to identify host resistance. This study defines A. trifolii as a significant but largely unknown contributor to severe root disease of subterranean clover in southern Australia. Finally, development and calibration of a new soil commercial DNA test not only enables field quantification of the disease, but development of appropriate breeding, selection and farm management strategies to reduce its impact.     

Effect of long-term changes in relative resource availability on dietary preference of grazing sheep for perennial ryegrass and white clover

Three replicate paddocks, each of 0·235 ha, containing adjacent monocultures of perennial ryegrass or white clover [50:50 by ground area, 6 cm sward surface height (SSH) at start of experiment] were continuously stocked with three yearling and four mature non-lactating, non-pregnant Scottish halfbred ewes for 12 weeks. Herbage intake, grazing behaviour and dietary selection were measured on seven occasions. Clover SSH declined rapidly over the first 5 weeks then stabilized at 1·2–1·6 cm, whereas perennial ryegrass SSH rose slightly initially, then declined gradually. Animals initially included proportionately c . 0·6 white clover in their diet but, by the end of the experiment, this had fallen to 0·3. Total daily herbage intake declined over the 12 weeks from 1·8 kg dry matter (DM) day^–1 at the start to 1·0 kg DM day^–1. Total grazing time increased from 561 min day^–1 to 649 min day^–1 at the end of the experiment. The results suggest that, despite overall herbage depletion and a greater depletion of white clover than perennial ryegrass as a result of the initial partial preference for white clover, the animals traded-off a reduced total intake and an increased grazing time in an attempt to maintain their initial preferred dietary composition.     

Yield and nitrogen concentration of above- and below-ground biomasses of red clover cultivars in pure stands and in mixtures with three grass species in northern Europe

Three red clover (Trifolium pratense) varieties differing in productivity and winter hardiness, Jokioinen, Betty and Ilte, were sown in a 2‐year (2003–2004) pot experiment in pure stands and mixtures with the grasses, timothy (Phleum pratense), tall fescue (Festuca arundinacea) and meadow fescue (Festuca pratensis). Grass growth dominated until fertilizer‐N, applied when the stands were sown, was depleted. Timothy was the least competitive of the grass species. Red clover variety Ilte produced the highest dry‐matter (DM) yields. Variety Betty yielded less, but allocated as much biomass to the root and stubble (soil‐bound) fraction as variety Ilte. Variety Jokioinen allocated least to the soil‐bound fraction. While the root structure and the starch concentration of the crown‐root area were similar in all varieties, the high ratio of soil‐bound: harvested fractions could be a key to the higher winter survival and higher DM yields of Betty under field conditions. At the end of the experiment, 3–5 g N pot^?1 (49–81 g m^?2) had been harvested and 0·7–1·5 g N pot^?1 (11–24 g m^?2) was left in the soil‐bound fraction, amounts depending on the red clover variety and grass mixture, with pure clover stands containing the highest N amounts. Because of the high N concentrations in the biomass of red clover, the proportion of red clover and conditions prevailing during canopy and root death in mixed stands are crucial for N mineralization and incorporation into new growth.     

Are seeding densities an opportunity to increase grain yield of winter wheat in a living mulch of white clover?

Optimum plant densities are a key to maximise yields in most crops. However, such information is often lacking for more environmentally sound cropping systems, such as living mulches (LM) for small grains. In 2004 and 2005, three trials were conducted in the Swiss Midlands on fields managed in accordance with the Swiss organic farming guidelines. The objective of the study was to determine whether seeding density of winter wheat (Triticum aestivum L.) is a relevant factor for determining grain yield in a white clover (Trifolium repens L.) living mulch. The winter wheat cv. Titlis was directly sown in wide spaced rows (0.375 m) at densities of 300 (LM300), 450 (LM450) or 600 (LM600) viable grains m⁻² in a white clover living mulch established at a seeding rate of 15 kg ha⁻¹. A bare soil control treatment with a wheat density of 450 viable grains m⁻² (BS450) was also included in the trials. Mean grain yields of LM300, LM450, and LM600 never reached the values observed in BS450. This was mainly due to a lower ear density, which, nevertheless, increased linearly with the seeding density within the living mulch in all trials, but the rate of increase depended on the environment. The decrease of the grain weight brought about by the increasing seeding density had only a marginal impact on the grain yield, which was increased from 1.31, 1.98, and 4.09 Mg ha⁻¹ (LM300) to 1.97, 2.64, and 4.75 Mg ha⁻¹ (LM600) for each of the three trials in the study. Significantly higher protein contents were observed for LM300 compared to the higher densities in the living mulch and to BS450. Our research showed that an increase of the seeding density is an effective mean to increase the grain yield in living mulch systems with white clover. However, it is likely that the control of the living mulch to reduce competition with the main crop is a more relevant factor.