小麦SSR分析体系的简化

摘要：SSR标记被广泛地用于小麦遗传育种研究，其技术流程主要包括基因组DNA提取、PCR扩增和PAGE凝胶电泳与染色。为了降低试验成本、缩短试验周期，建立一套经济、快捷的SSR分析体系，（1）比较了不同提取方法提取的DNA和从小麦幼苗叶片、种子和种胚中提取DNA的效果；（2）比较了不同的PCR反应体系；（3）比较了几种PAGE胶银染方法。结果证明，以简化CTAB法提取的小麦种胚DNA为模板，采用10µLPCR反应体系和快速银染法染色可以得到与原方法同样的效果，节省了50%的试验时间和50%的试验成本。     

一个斯卑尔脱小麦早熟突变体抽穗期的遗传分析与分子标记初步定位

研究以斯卑尔脱小麦(Triticum spelta)Hubel及其早熟诱变系2463为材料,采用主-多基因混合模型多世代联合分析发现,抽穗期性状由两个主基因控制,其遗传力为44%～81%.温室春化和光周期处理实验结果表明,经过春化处理后,与Hubel相比,其早熟诱变系在长日照条件下的抽穗期提前10d左右,表现为显著差异.利用F2分离群体进行的分子标记定位结果表明,在2D染色体的短臂上存在一个主效QTL,与标记Xwmc112紧密连锁,能解释表型变异的16%,与前人定位的光周期相关QTL/基因(Ppd1)位于同一区域.     

小麦优异品系遗传构成的简单重复序列(SSR)分析

为了探讨优良品种形成的遗传学基础,本研究利用简单重复序列(SSR)标记分析了近期育成的3个丰产小麦(Triticum aestivum L.)品系(百农金光588、百农AK58-18和百农T5)及其姊妹系(百农0487和百农高光3709)的遗传构成.结果表明,2个杂交组合(周麦18/百农AK58和周麦16/温麦8号//百农160)中亲本对不同姊妹系的遗传贡献率均具有较大差异,其中百农AK58对百农金光588、百农AK58-18和百农0487的遗传贡献率分别为51.2％、57.0％和54.0％,高于亲本周麦18的贡献率;百农160对百农高光3709和百农T5的遗传贡献率分别为41.9％和36.4％,高于另外2个亲本(周麦16和温麦8号)的贡献率.在A、B、D基因组及21对染色体上,衍生后代对不同亲本遗传物质的继承率也表现丰富的多样性.百农金光588、百农AK58-18和百农T5分别具有109、36和55个不同于其他姊妹系的SSR特异位点,分别形成了11、3和3个特异染色体区段.这些基因组区段上存在许多与产量和抗病等重要农艺性状相关的基因和QTL,对提高小麦丰产性可能起了重要作用.     

小麦抗叶锈病基因Lr2c的SSR标记

选取抗叶锈病基因位于2D染色体上的TcLr2c等7个小麦（Triticum aestivum）近等基因系、感病亲本Thatcher及215株TcLr2c与Thatcher杂交F2代为材料,研究抗叶锈病基因Lr2c SSR分子标记。从筛选的29对位于小麦2D染色体的SSR引物中获得4对能够揭示Lr2c多态性的分子标记,通过215株TcLr2c × Thatcher F2群体验证,结果表明Xgwm261和Xgwm296与Lr2c紧密连锁,其距目的基因的遗传距离分别为1.9和3.6 cM,可用于小麦抗叶锈病分子辅助育种。     

小麦RIL群体SSR标记偏分离的遗传分析

以普通小麦3338与斯卑尔脱小麦Altgold杂交得到的F6代重组近交系群体为材料，筛选出334个（271个SSR和63个EST-SSR标记）多态性标记，以此为基础构建了一个包含287个分子标记的遗传图谱。对这些多态性标记进行偏分离分析发现82个表现为偏分离（P＜0.05），其中70个可以定位到遗传连锁图谱中。这些偏分离标记中有33个标记位点偏向母本3338，占40.2%，49个标记位点偏向父本Altgold，占59.8%。这些偏分离标记在图谱上的分布有两种：成簇分布和孤立位点的偏分离。在5条不同染色体上发现6个偏分离热点区域，这些偏分离热点区域的形成可能与配子体选择有关。     

14份小麦种质资源抗麦长管蚜遗传多样性的SSR分析

利用SSR分子标记在分子水平探讨小麦种质资源抗麦长管蚜的遗传多样性,为高效节本型和环境友好型的抗蚜育种的研究和利用提供理论依据和技术支持。结果表明：在小麦的A、B和D 3组同源染色体组上的175对SSR分子标记鉴定出了有多态性的32对引物,其中在D同源组中小麦抗蚜性的遗传多样性较高,同时在21对染色体中7D染色体上遗传...     

DNA分子标记技术及其在小麦育种及遗传研究中的应用

本文介绍了几种常用的DNA分子标记,如RFLP、RAPD、AFLP、SSR、STS、SNP等,并简要综述了分子标记技术在小麦遗传育种研究中的应用现状,包括基因标记与定位、遗传图谱构建、外源染色体鉴定与标记、种质资源鉴定和辅助育种等。     

我国部分地区小麦叶锈菌遗传多样性的SSR分析

小麦叶锈菌(Puccinia triticina)引起的小麦叶锈病是小麦上的重要病害。为了解小麦叶锈菌遗传多样性及其亲缘关系,本研究利用简单重复序列(simple sequence repeat,SSR)标记技术对2009年采自河北、河南、山东、四川4省的小麦叶锈菌株进行SSR分析。小麦叶锈菌的观察等位基因数(Na)为1.75,有效等位基因数(Ne)为1.40,Shannon信息指数(I)为0.35,Nei’s基因多样性指数(H)为0.23,多态性百分率为75.29%,其中河北、河南和山东的叶锈菌群体遗传多样性水平高于四川群体。聚类分析表明,在相似系数0.96处4个群体聚为2组,河南、山东及四川群体聚为一组,河北群体自成一组,其中河南和山东群体亲缘关系最近。小麦叶锈菌具有一定的遗传变异,群体间遗传变异占总变异的8.93%,群体内遗传变异占总变异的91.07%。小麦叶锈菌群体间每代迁移数Nm为6.10。小麦叶锈菌遗传多样性丰富,群体间遗传相似性较高,亲缘关系与地理分布具有一定相关性。群体内遗传变异是群体遗传变异的主要来源。本研究说明群体间存在广泛的菌源交流,为明确小麦叶锈病流行区系和叶锈菌传播路线提供了基础资料。     

小麦-柔软滨麦草易位系M853-4抗条锈病基因的遗传分析和SSR标记定位

用中国小麦条锈菌(Puccinia stniform f.sp.stritici)流行小种条中29号、条中30号、条中31号和水源11-11生理小种,对小麦(Triticum aestivum)-柔软滨麦草(Elymus mollis)易位系M853-4和铭贤169的杂交后代进行苗期抗条锈性遗传分析.结果表明,易位系M853-4对条中29号、条中30号和条中31号的抗病性均由1对显性核基因控制;对水源11-11的抗病性由1对显性和1对隐性基因共同控制.将控制条中31号抗病性的基因暂时命名为YrElm4.以接种条中31号的F2正交群体为研究对象,应用BSA法进行了SSR分析.从320对SSR引物组合中筛选到3个与主效抗病基因YrE1m4连锁的多态性微卫星标记,它们分别是Xwmc654、Xgwm304和Xgwm129,与YrE1m4的遗传距离依次为5.8、7.1和10.3 cM,并将YrE1m4定位于小麦5AS染色体,这3个标记可用于分子标记辅助育种.     

DNA分子标记技术在小麦遗传育种中的应用综述

在简述DNA分子标记的种类和特点的基础上,综述了分子标记技术在小麦遗传育种研究中主要应用于遗传图谱构建、基因标记和定位、品种鉴定和指纹图谱绘制、物种亲缘关系和遗传多样性研究及分子标记辅助育种等方面的现状,并对分子标记技术的发展和应用前景进行了展望。     

Effectiveness of Zinc and Gypsum Application Against Cadmium Toxicity and Accumulation in Wheat (Triticum aestivum L.)

Cadmium (Cd) is a nonessential and toxic element because it inhibits the growth and development of plants and is dangerous for end consumer. It enters in the human food chain through food crops. Application of plant nutrients such as zinc (Zn) and gypsum is a viable and cheap strategy to minimize its accumulation in edible plant portions. Therefore, this investigation was conducted to determine the effectiveness of Zn and gypsum against Cd accumulation in wheat. The results showed that Cd toxicity considerably decreased the plant growth, physiological activities, and yield attributes and increased the Cd accumulation in root, shoot, and grain, while application of Zn and gypsum remarkably increased the growth and yield and decreased the Cd accumulation in plant parts in Cd-contaminated soil. The results also depicted that application of Zn showed better results as compared to gypsum. In conclusion, we can say that application of Zn and gypsum remarkably ameliorated the Cd toxicity and decreased its accumulation in wheat, grown in Cd-contaminated soil.     

小麦显性多子房基因的RAPD标记

利用BSA法，以显性多子房小麦（rriticum aestivum）与普通小麦（T.aestivum）品种西农1376进行杂交，从F2分离世代建立多子房性状基因池和单子房基因池，采用RAPD-PCR方法，以440个随机引物筛选单、多子房的混合群体及其双亲。发现10个引物的扩增产物在近等基因系间表现出特异性，表现为多子房亲本及其混合群体有带，单子房亲本及其混合群体无带；6个引物扩增产物表现出的特异性相反，单子房有带，多子房无带。经过重复实验和群体单株验证，引物S236在多子房亲本、混合群体及其群体单株中都表现出稳定的多态性，可以作为多子房显性基因的分子标记。     

Lox基因RNAi转基因小麦后代株系种子耐储藏性分析

小麦(Triticum aestivum)种子中脂氧合酶(lipoxygenase,LOX)可催化不饱和脂肪酸的氧化作用,在种子储藏期间产生氢过氧化物,导致小麦种子在储藏过程中种子活力和营养品质降低.因此,选育种子LOX活性低的小麦品种是小麦遗传改良的任务之一.为了获得种子脂氧合酶活性显著降低、外源Lox RNAi构件(Lox geneRNAiconstruct,Loxi)能稳定遗传且种子耐储藏性增强的转基因小麦新种质,本研究以小麦西农889、闫麦8911和陕优225转Loxi稳定遗传的TF7代转基因株系为材料,对转基因种子中Lox基因家族进行了半定量RT-PCR分析,结果表明,外源Loxi可有效抑制小麦籽粒中小麦脂氧合酶1(Triticum aestivum lipoxygenase Ⅰ,TaLox1)基因的表达,但小麦脂氧合酶2(Triticum aestivum lipoxygenase 2,TaLox2)和小麦脂氧合酶3(Triticum aestivum lipoxygenase 3,TaLox3)基因的表达量不变;对3个亲本及16个TF7代转基因株系的种子脂氧合酶活性检测,结果显示,与其亲本相比,转基因株系种子脂氧合酶活性显著降低;对人工加速老化处理7和14 d的转基因和对照种子分析发现,随着老化天数的增加种子活力降低,但转基因株系种子活力均显著高于其亲本材料;人工加速老化后脂肪酸组分和含量测定分析表明,转基因株系脂肪酸比其亲本材料氧化损失少.以上结果说明,转基因株系中Lox基因的RNAi可有效抑制其种子LOX活性,使转基因小麦后代种子耐储藏性增强;筛选鉴定得到的种子LOX活性显著降低且耐储藏性增强的转基因小麦株系可为小麦育种提供新的材料.     

低温诱导的位点特异性重组植物表达载体构建及小麦转化

选择标记基因的剔除是转基因植物商品化种植的重要基础.为建立小麦(Triticum aestivum)低温诱导的位点特异性标记基因删除体系,本研究以小麦西农928基因组DNA为模板,通过PCR扩增获得小麦低温诱导表达wcs120启动子,序列分析表明,该序列与GenBank中序列AF031235相比有一段21 bp插入,与AY493570.1序列完全一致.以含有CinH/RS2单向位点特异性重组系统的植物表达载体pXL5513为框架,成功构建了包含有抗旱相关性基因PYL5(pyrabactin-resistance like gene 5)表达框的低温诱导的位点特异性重组植物表达载体pXL5513-fwcs-RDA;并对小麦绵阳19(M19)幼胚愈伤组织进行基因枪转化,1 745块愈伤组织经筛选分化共获得9株转基因植株,经PYL5基因特异引物PCR检测,6株为阳性植株,低温春化处理后经删除引物PCR检测,6株阳性植株初步证实成功删除了选择标记基因.本研究为利用低温诱导的位点特异性重组系统进行小麦无标记基因转化奠定了基础.     

Interactive Effects of Salinity Stress and Zn Availability on Physiological Properties,Antioxidant Activity,and Micronutrients Content of Wheat (Triticum aestivum) Plants

In order to evaluate the combined effects of salinity stress and Zinc (Zn) applications on wheat growth, an experiment was conducted with the treatments included four Zn levels (0, 10, 20, and 40 mg/kg of dried soil) and three levels of sodium chloride (NaCl) salinity of irrigation water (0, 100, and 200 mM NaCl). Salinity caused a significant reduction in chlorophyll a and b content. The activity of catalase (CAT), superoxide dismutase (SOD), and glutathione reductase (GR) was enhanced as salinity level was increased. The significant enhancement in concentration of sodium due to the salinity was alleviated by Zn addition. Potassium content was increased by Zn treatments. A substantial increase was observed in leaf Zn concentration as the applied level of Zn was increased. Overall, these results indicate some positive and negative interactions of salinity and Zn application, which could be helpful in management of the saline soil and water.     

小麦T型细胞质雄性不育恢复基因Rf1和f4的SSR标记分析

为进一步探讨小麦(Triticum aestivum L.)T型细胞质雄性不育(T-CMS)育性恢复的遗传机理,并为利用T型不育系选育强优势杂交小麦分子辅助育种提供理论与技术支撑,本研究以小麦ms(S)矮抗58/R113的F2代分离群体中的极端可育株和极端不育株分别建立恢复池和不育池,采用分布于小麦第一染色体群(染色体1A、1B和1D)及第六染色体群(染色体6A、6B和6D)上的196对SSR引物进行扩增筛选.结果表明,(1)位于1AS染色体上的3个SSR标记和位于6BS染色体上的4个SSR标记均在亲本和基因池间扩增出了稳定的多态性差异条带;(2)定位群体验证结果表明,恢复基因Rf1与1AS染色体上Xgwm136、Xgpw7062和Xgdm33标记的遗传距离分别为4.8、9.6和13.7 cM,3个标记与Rf1之间的顺序依次为Xgdm33、Xgwm136、f1、Xgpw7062; (3)恢复基因Rf4与6BS染色体上的Xgpw1079、Xgwm193、Xgpw7011和Xgwm508标记的遗传距离分别为3.4、6.8、13.7和21.5 cM,4个标记与Rf4之间的顺序依次为Xgpw 011、Xgpw1079、Rf4、Xgwm19和Xgwm508.研究还表明,T-CMS恢复系R113的育性是由Rf1和f4两对主效恢复基因和多对微效基因共同控制的,筛选的上述7个SSR标记可直接用于T型或者类似T型,如S型杂交小麦分子标记辅助育种,可有效提高对应恢复系的选择效率.     

Soil and Plant Responses to the Application of Ascophyllum nodosum Extract to No-Till Wheat (Triticum aestivum L.)

A field trial consisting of four granule formulation doses and five liquid formulation sprays of a seaweed extract from Ascophyllum nodosum commercially known as Biovita, along with the recommended dose of nitrogen (N)–phosphorus (P)–potassium (K), was conducted during 2008 and 2009 in BHU, Varanasi, India, to evaluate its effect on wheat (var. HUW 468) under a no-tillage system. Among the granule doses, the 10 kg ha^?1 basal application and the two liquid sprays of 500 cm³ ha^?1 each at 25 and 50 days after sowing significantly improved the performance of wheat. On an average under these two treatments, the greatest grain and straw yields were observed were 3454.5, 3446.5 and 5187.5, 5220.0 kg ha^?1, respectively. The greatest protein content was found when further high doses of extract were applied. A faster decomposition of the paddy residue was also observed as indicated by an earlier decline in carbon (C)/N ratio of the soil in the treated plots.     

In-Season Prediction of Nitrogen Use Efficiency and Grain Protein in Winter Wheat (Triticum aestivum L.)

In a 3-year study, grain yield, nitrogen use efficiency (NUE), and grain protein (GP) were evaluated as a function of rate and timing of nitrogen (N) fertilizer application. Linear models that included preplant N, normalized difference vegetation index (NDVI), cumulative rainfall, and average air temperature from planting to sensing (T-avg) were evaluated to predict NUE and GP in winter wheat. GreenSeeker readings were collected at Feekes (F) 3, 4, 5, and 7 growth stages. Combined with rainfall and/or T-avg, NDVI alone was not correlated with NUE. However, NDVI and rainfall explained 45% (r² = 0.45) of the variability in GP at F7 growth stage. Preplant N, NDVI, rainfall and growing degree days (GDD) combined explained 76% (r² = 0.76) of the variability in GP at F3. Mid-season climatic data improved the prediction of GP and should therefore be considered for refining fertilizer recommendations when GP levels are expected to be low.     

Copper induced iron deficiency in wheat (Triticum aestivum L)

Abstract

A pot experiment was conducted to study the interaction effects of phosphorus and copper on wheat. The soils used were calcareous loamy sand (ls) and non calcareous sandy loam (sl). Four levels of Cu (0, 5, 10 and 20?mg Cu kg^?1 soil) and six levels of P (0, 25, 50, 100, 200 and 400?mg P kg^?1 soil) were applied in all possible combinations with three replications. Soil pH decreased with Cu application while Olsen P increased with P application in both soils. Growth and yield of wheat improved significantly with graded levels of applied P. However, when any level of P was combined with 20?mg Cu kg^?1 soil, severe iron chlorosis of leaves, a drastic reduction in growth and chlorophyll content was observed in calcareous ls only. The results indicated that it was Cu and not P that induced Fe deficiency in wheat grown in alkaline calcareous soil and the Cu requirement of the crop seemed to be much lower in the calcareous ls. Root dry matter, grain and straw yield decreased with increasing levels of applied Cu in ls but in sl maximum increase of 62.5, 74.3 and 63.7 per cent in root, grain and straw yield was observed with a combined application of 400?mg P and 5?mg Cu kg^?1 soil over control. Accumulation of Cu in roots decreased the Fe absorption by roots which indicated that Fe chlorosis of wheat leaves is expected when Cu: Fe concentration ratio in root is > 0.30.