小麦条锈菌诱导性小麦cDNA文库的构建及其质量评价

小麦条锈病是由小麦条锈菌(Puccinia striiformis f.sp.tritici)侵染引起的世界性气传叶部病害之一,在我国发生尤为普遍而严重,是小麦(Triticum aestivum L.)生产上最重要病害,克隆并研究小麦抗条锈病相关基因的生物学功能具有重要的应用价值和现实意义.在本研究中用中国当前毒性最强优势条锈菌小种CY32侵染诱导的小麦抗条锈病基因Yr5近等基因品系Taichung29*6/Yr5为试材,构建非亲和条锈菌小种侵染诱导的小麦cDNA文库.研究结果表明所获得的原始文库滴度0.9x106 pfu/mL,扩增文库滴度1.0x109pfu/mL,重组率98%,插入片段在0.5 kb到3.0 kb之间,多在1.0 kb左右.因此该文库可用于基因克隆与分析,为研究小麦抗条锈病相关基因提供条件.     

33个小麦品种(系)抗叶锈基因Lr19分子检测

由小麦叶锈菌(Puccinia triticinia)引起的小麦叶锈病在世界各地产麦区均有发生。利用抗病品种是防治该病害最经济、安全、有效的方法。小麦抗叶锈病基因Lr19是一个十分有效的抗叶锈性基因,自1966年首次将该基因从长穗偃麦草(Agropyron elongatum)转到普通小麦中,至今仍是一个应用潜力很大的抗病基因。本研究利用以PCR为基础的STS技术对以春小麦Thatcher为背景的50个近等基因系材料和TcLr19与Thatcher杂交F2小麦进行检测,并对33个小麦品种进行分子标记分析鉴定,结果如下:(1)对以春小麦Thatcher为背景的50个近等基因系材料和TcLr19×ThatcherF2代小麦进行PCR-STS检测,扩增结果表明在50个近等基因系材料中仅有TcLr19中出现一条130bp的DNA条带,STSLr19130标记在其他49个近等基因系材料中未检测到Lr19基因;F2代中表现感病的植株没有130bp的DNA片段,表现抗病的植株有130bp的DNA条带;重复两次结果相同,进一步证明了与小麦抗叶锈基因Lr19共分离的STS标记的稳定可靠。(2)利用以PCR为基础的STSLr19...     

河北省又取得一批农药植保类科研成果

<正>(接上期)8.小麦抗叶锈病相关基因的克隆单位名称:河北农业大学评价单位名称:河北省教育厅叶锈病是小麦高产、稳产的主要限制因素,选育和利用抗病品种是控制小麦叶锈病最为经济有效的措施。但由于病菌新毒性基因的不断产生以及致病性分化加快,新的具有更强致病能力的优势生理小种不断出现,致使已有的一些抗叶锈基因逐渐丧失抗性。该项目以含有小麦抗叶锈基因Lr41的小麦品种KS90WGRC10为材料,根据从抑制差减杂交文库中     

CIMMYT小麦材料的苗期和成株抗叶锈病鉴定

选择来自CIMMYT的103个小麦品种(系)及35个含有已知抗叶锈基因的对照品种,苗期接种17个中国小麦叶锈菌小种以鉴定这些品种中可能含有的抗叶锈病基因, 并于2008-2009和2009-2010连续2年对这些材料进行成株抗叶锈病鉴定。通过苗期鉴定结合系谱分析和分子检测,在46个品种中共鉴定出Lr26、Lr34、Lr42和Lr47等4个抗叶锈病基因,其中9个品种携带Lr26基因,28个品种含有成株抗叶锈病基因Lr34基因,Lr42可能存在于11个品种中,还有2个材料可能含有Lr47,其他57个品种(系)对供试的15个小种多数表现为高抗,没有鉴定出已知的抗叶锈病基因。通过两年的田间抗叶锈病鉴定共筛选出46个表现慢锈的品种。苗期和田间结果表明,CIMMYT材料中含有丰富的对我国叶锈菌小种有效的苗期和成株抗叶锈基因,这些材料均可应用于我国小麦的抗叶锈病育种。     

河北省又取得一批农药植保类科研成果

<正>9.小麦抗叶锈病基因的分子标记研究单位名称:河北农业大学评价单位名称:河北省科技成果转化服务中心利用基因推导、抗病性遗传分析以及SSR分子标记技术对具有良好的综合农艺性状,但其所携带的抗叶锈基因尚不清楚的小麦品种毕麦16和Muu中的抗叶锈基因进行遗传特性研究;同时利用SRAP、c DNA-AFLP技术对小麦抗叶锈病基因Lr19和Lr41进行分子标记研究,结果如下:(1)毕麦16对叶锈菌生理小种FHTT的抗性由1对     

70份国外小麦品种(系)的苗期和成株期抗叶锈病鉴定

小麦叶锈病是小麦生产中的重要病害之一,培育持久抗病品种是最经济、有效和环保的方法。本研究用19个不同毒力的叶锈菌小种苗期接种70份国外引进小麦品种(系)及36个已知抗叶锈病基因的载体品种进行抗性鉴定,同时在2016—2017年度分别于河北保定和河南周口对70份国外引进品种进行田间抗叶锈性鉴定。为进一步检测材料中所携带的苗期和成株抗叶锈病基因,利用12个与已知基因紧密连锁的分子标记进行检测,综合基因推导、系谱分析和分子标记检测的结果,在33份材料中鉴定出15个抗叶锈病基因,包括Lr1、Lr2a、Lr26、Lr3ka、Lr11、Lr17、Lr30、Lr10、Lr14a、Lr2b、Lr13、Lr15、Lr21、Lr44和Lr45,田间鉴定筛选出39份品种表现慢锈性。苗期和田间表现表明,国外品种中含有丰富的对我国叶锈菌小种有效的苗期和成株期抗叶锈病基因,可作为小麦抗叶锈病抗源在抗病育种中加以利用。     

山西省不同生态区小麦叶锈菌毒性研究

利用已知抗叶锈病基因的小麦近等基因系 (或单基因系 )作鉴别寄主 ,监测来自山西省 3个不同小麦生态区 11个县 (市 )的 92份叶锈标样。在发现的 2 7个致病类型 (毒性基因组合 )中 ,TRK ,TRT ,PHT ,THT出现频率分别为 19 6% ,9 8% ,6 5 % ,6 5 % ,为优势致病类型。对叶锈菌群体毒性基因频率分析结果表明 ,毒性基因V19,V2 4 ,V38的出现频率较低 ,分别为5 4 % ,16 5 %和 0 ,其对应的抗性基因可视为山西省小麦叶锈菌的有效抗病基因     

谷子十里香抗锈抑制消减杂交文库构建及初步分析

为了研究十里香抗锈分子机理及其调控机制。以谷子抗锈十里香接种12,24,48,72,96 h叶片为材料,利用抑制差减杂交技术,构建了谷锈菌诱导的SSH文库,筛选十里香接种与未接种锈菌差异表达的基因片段,通过Gen-Bank进行同源比对,对差异表达基因进行功能注释,并利用荧光定量PCR技术对部分差异表达片段进行表达分析。随机挑取差减文库中阳性克隆测序,共获得368个EST序列,插入片段大小为200~750 bp,通过网上GenBank非冗余数据库比对分析,发现其中32个EST与抗病相关。对与抗病相关的EST分析,推测WRKY转录因子、MAPK信号途径、钙信号途径、谷胱甘肽-S-转移酶、细胞色素P450、病程相关蛋白等可能参与了十里香与谷锈菌非亲和互作。进一步利用荧光定量PCR技术对SSH文库中4个基因做了表达分析,结果表明这些基因均受锈菌诱导表达。通过构建十里香受锈菌诱导的SSH文库,初步明确了十里香参与抗锈相关的基因,为下步谷子抗锈分子育种奠定基础。     

TaRanGAP2在小麦抵抗叶锈菌侵染中的作用

为研究TaRanGAP2在小麦抵抗叶锈菌侵染的HR诱发中的作用,阐明小麦抗叶锈病分子机制,为小麦抗叶锈病育种给予分子水平的指导.以小麦近等基因系TcLr26及其轮回亲本Tc分别与叶锈菌生理小种260组成不亲和组合(TcLr26×260)及亲和组合(Tc×260),生物信息学分析发现,TaRanGAP2的CDS全长为16...     

小麦NBS类抗病相关基因片段RGA-A的初步研究

利用同源序列法分离小麦抗病相关基因同源序列。根据已经克隆的抗病基因保守结构NBS区设计引物,采用RT-PCR方法对小麦抗叶锈病近等基因系材料TcLr24进行扩增。获得了一条通读的525 bp条带RGA-A,通过BLASTp比较,序列中含有典型的NBS保守结构域,与很多已知植物抗病基因的功能相应区域一致,编码的蛋白与大麦中抗性蛋白亲缘关系较近。半定量RT-PCR分析表明,RGA-A受叶锈菌诱导表达,表明该基因在小麦叶片中与抗叶锈性相关。该NBS类抗病基因相关片段的获得为研究小麦抗病基因奠定了基础。     

28个小麦微核心种质抗叶锈性分析

选取在成株期表现高、中、低抗叶锈的28个小麦微核心种质,利用39个以Thatcher为背景的近等基因系(或单基因系)作为已知基因的鉴别寄主,接种8个小麦叶锈菌致病型进行苗期抗叶锈基因推导,结合成株期抗病鉴定,初步明确了这些品种(系)的抗性和可能携带的抗病基因。利用19个与Lr基因紧密连锁或共分离的分子标记,对28个微核心种质进行抗叶锈病基因的进一步鉴定,推测新克旱9号可能含有Lr17、Lr2b、Lr14a和Lr33;兴义4号可能含有Lr26、Lr36和Lr37;紫皮可能含有Lr2b和Lr34;大白皮含有Lr1;毕红穗含有Lr1、Lr10和Lr34;中优9507含有Lr10;小白麦、红粒当年老、老麦、蝉不吱、苏麦3号和车锏子含有Lr1和Lr34;红花早可能含有Lr1、Lr34、Lr14a和Lr2b;江西早、泡子麦、三月黄、有芒扫谷旦、阜阳红、成都光头和酱麦可能含有Lr34;敦化春麦和甘肃96可能含有Lr28;欧柔可能含有Lr34、Lr16、Lr11、Lr3bg和Lr33;此外,新克旱9号、兴义4号、红花早、红粒当年老、欧柔、有芒扫谷旦、成都光头、甘肃96、小红皮、定兴寨、中优9507和红冬麦中可能含有未知抗病基因;在这28份种质中,不含Lr9、Lr19、Lr20、Lr21、Lr24、Lr29、Lr35、Lr38和Lr47基因。研究结果表明,测试的微核心种质中含有比较丰富的抗叶锈病基因,可为育种提供丰富的抗源。     

56个小麦品种（系）的苗期和成株抗叶锈鉴定

为了研究中国小麦品种中所携带的抗叶锈基因,对56个小麦品种（系）进行苗期接种推导其中所含有的抗叶锈基因,同时连续2年对供试材料进行田间成株抗叶锈鉴定。通过苗期基因推导结合分子标记辅助检测,结果表明,在36个小麦品种中共鉴定出Lr26、Lr34、Lr1、Lr2a、Lr11、Lr20、Lr30、Lr33和Lr44等9个抗叶锈基因,其中28个品种含有Lr26,Lr1和Lr20分别存在于6个品种中,4个品种含有Lr30,Lr11和Lr44各存在于2个品种中,Lr2a、Lr33和Lr34各自在1个品种中出现。经过2年的田间抗叶锈鉴定共筛选出46个慢锈品种。筛选到的这些苗期和成株抗病品种均可用于小麦持久抗叶锈品种的培育。     

土耳其8个小麦品种农艺性状及抗叶锈性分析

为确定8个来自土耳其的普通小麦品种在我国的应用前景,对其进行全生育期农艺性状观察,并利用44个以Thatcher为背景的近等基因系(单基因系)作为已知基因的鉴别寄主,接种8个小麦叶锈菌致病型进行苗期抗叶锈基因推导,结合成株期抗病鉴定,初步明确了这些品种(系)的抗性和可能携带的抗病基因。利用20个与Lr基因紧密连锁或共分离的分子标记,对8个土耳其小麦品种进行抗叶锈病基因的进一步鉴定。推测YJ000900中可能含有Lr1、Lr3、Lr17、Lr20;YJ000906中可能含有Lr1、Lr17、Lr20;YJ000901、YJ000902、YJ000904、YJ000905、YJ000907中可能含有Lr1;8个材料中均不含Lr9、Lr19、Lr20、Lr21、Lr24、Lr26、Lr28、Lr29、Lr34、Lr35、Lr37、Lr38和Lr47基因。结果表明,来自土耳其的8个小麦材料具有较差的抗叶锈性、抗寒和抗倒伏能力,而且产量低,不适宜于大规模推广种植,也不能作为我国小麦抗叶锈的抗源使用。     

小麦抗叶锈病基因Lr45的SSR分子标记

用定位于2A染色体的59对SSR、EST-SSR引物,对小麦抗叶锈病基因Lr45进行分子标记,共筛选出11对揭示TcLr45多态性的引物。用157株F₂抗感群体对这11对引物进一步检测,得到4个与Lr45共分离的SSR标记(Xgwm95、Xgwm47、Xgwm372和Xgwm122)。将经PAGE检测的标记Xgwm95的抗感差异带及Xgwm47的抗性片段进行克隆测序发现其中含有微卫星序列,且均为二核苷酸重复。Xgwm372和Xgwm122经琼脂糖凝胶电泳发现与Lr45的供体黑麦有共同的标记片段,可直接用于分子标记辅助选择。     

Identification of a molecular marker linked to an Agropyron elongatum-derived gene Lr19 for leaf rust resistance in wheat

The leaf rust resistance gene Lr19, transferred from Agropyron elongatum into wheat (Triticum aestivum L.) imparts resistance to all pathotypes of leaf rust (Puccinia recondita f.sp. tritici) in South‐east Asia. A segregating F₂ population from a cross between the leaf rust resistant parent ‘HW 2046’ carrying Lr19 and a susceptible parent ‘Agra Local’ was screened in the phytotron against a virulent pathotype 77‐5 of leaf rust with the objective of identifying the molecular markers linked to Lr19. The gene was first tagged with a randomly amplified polymorphic DNA (RAPD) marker S73₇₂₈. The RAPD marker linked to the gene Lr19 which mapped at 6.4 ± 0.035 cM distance, was converted to a sequence characterized amplified region (SCAR) marker. The SCAR marker (SCS73₇₁₉) was specific to Lr19 and was not amplified in the near‐isogenic lines (NILs) carrying other equally effective alien genes Lr9, Lr28 and Lr32 enabling breeders to pyramid Lr19 with these genes.     

A leaf rust resistance gene, different from Lr34, associated with leaf tip necrosis in wheat

The gene Lr34 has contributed to durable resistance to leaf rust caused by Puccinia triticina in wheat worldwide. The closely associated leaf tip necrosis is generally used as the gene's marker. Lr34 has been postulated in many Indian bread wheat cultivars including ‘C 306’, based on the associated leaf tip necrosis and a few other field and glasshouse observations. The present study showed monogenic control of adult‐plant resistance in ‘C 306’ to leaf rust pathotype 77‐5 (121R63‐1). The F₂ segregation in the crosses between ‘C 306’ and the two known carriers of Lr34, ‘Line 897’ and ‘Jupateco 73’‘R’ fitted a digenic ratio. The F₃ families derived from the susceptible F₂ segregants were true breeding for susceptibility, proving the absence of Lr34 in ‘C 306’. The cross between ‘Line 897’ and ‘Jupateco 73’‘R’ did not segregate for susceptibility. Resistance in the cross ‘Agra Local’ (susceptible) × ‘C 306’ was associated with leaf tip necrosis, showing that the leaf rust resistance gene in ‘C 306’ was associated with leaf tip necrosis, but was different from Lr34. This gene is being temporarily designated as Lr‘C 306’. Hence, leaf tip necrosis cannot be considered as an exclusive marker for selecting Lr34 in wheat improvement.     

Resistance to leaf rust in cultivars of bread wheat and durum wheat grown in Spain

A set of bread wheat and durum wheat cultivars adapted to Spanish conditions was tested for resistance against leaf rust caused by different pathotypes of Puccinia triticina in field trials and in growth chamber studies. Lower levels of resistance were found in durum wheat than in bread wheat. The most frequent Lr genes found in bread wheat were Lr1, Lr10, Lr13, Lr20, Lr26 and Lr28. In durum wheat, additional resistance genes that differed from the known Lr genes were identified. The level of partial resistance to leaf rust was in general low, although significant levels were identified in some bread wheat and durum wheat cultivars.     

‘CPAN 1842’: a new source of adult plant leaf rust resistance in bread wheat

There is worldwide interest in adult plant resistance (APR) because of greater durability of APR to the cereal rusts. Peruvian bread wheat genotype ‘CPAN (Coordinated Project Accession Number) 1842’ (LM 50–53) has shown leaf rust resistance in disease screening nurseries since its introduction in 1977. However, it is susceptible at the seedling stage to several Puccinia triticina (Pt) pathotypes including the widely prevalent 77‐5 (121R63‐1) that infects bread wheat. Inheritance studies showed that CPAN 1842 carried a dominant gene for APR to pathotype 77‐5, which was different from Lr12, Lr13, Lr22a, Lr34, Lr35, Lr37, Lr46, Lr48, Lr49 and Lr68, based on the tests of allelism; and from Lr67, based on genotyping with the closely linked SSR marker cfd71. This gene should also be different from Lr22b as the latter is totally ineffective against pathotype 77‐5. CPAN 1842 therefore appears to be a new promising source of leaf rust resistance. Also having resistance to stem rust and stripe rust, this line can contribute to breeding for multiple rust resistances in wheat.     

Development and Validation of SCAR Markers Co-Segregating with an Agropyron Elongatum Derived Leaf Rust Resistance Gene Lr24 in Wheat

Summary An Agropyron elongatum-derived leaf rust resistance gene Lr24 located on chromosome 3DL of wheat was tagged with six random amplified polymorphic DNA (RAPD) markers which co-segregated with the gene. The markers were identified in homozygous resistant F₂ plants taken from a population segregating for leaf rust resistance generated from a cross between two near-isogenic lines (NILs) differing only for Lr24. Phenotyping was done by inoculating the plants with pathotype 77-5 of Puccinia triticina. To enable gene-specific selection, three RAPD markers (S1302₆₀₉, S1326₆₁₅ and OPAB-1₃₈₈) were successfully converted to polymorphic sequence characterized amplified region (SCAR) markers, amplifying only the critical DNA fragments co-segregating with Lr24. The SCAR markers were validated for specificity to the gene Lr24 in wheat NILs possessing Lr24 in 10 additional genetic backgrounds including the Thatcher NIL, but not to 43 Thatcher NILs possessing designated leaf rust resistance genes other than Lr24. This indicated the potential usefulness of these SCAR markers in marker assisted selection (MAS) and for pyramiding leaf rust resistance genes in wheat.