条形柄锈菌34号生理小种的分子标记

条形柄锈菌Puccinia striiformis f. sp. tritici 34号生理小种(CYR34)是目前我国毒性谱最宽、毒性最强的生理小种,对小麦生产和抗病品种选育造成了极大的影响。本研究采用RAPD-SCAR分子标记技术,从300条RAPD随机引物中筛选到CYR34的特异引物,通过特异性片段回收、克隆和测序(GenBank登录号为OL907303),依据序列设计出了S2008F34/S2008R34特异性引物,能够从CYR34及接种CYR34的小麦发病叶片总DNA中都扩增出417 bp的目标片段。采用该特异性引物检测2021年陕西渭南、咸阳和宝鸡地区小麦条锈菌CYR34的流行频率分别为8.6%、6.0%和10.8%。该项研究为小麦条锈菌CYR34号生理小种的快速检测提供了技术支撑。     

小麦条锈菌条中31号生理小种SCAR检测标记的建立

建立小麦条锈菌Pucciniastriiformisf.sp.tritici生理小种的快速分子检测技术对我国小麦条锈病的监测和防治策略的制定具有重要价值,本文首次报道了利用SCAR—PCR技术进行条锈菌生理小种分子检测的方法。通过对我国目前主要优势小种条中31号RAPD片段的规模筛选,在对特异片段回收、克隆、测序的基础上,设计特异PCR引物,成功获得了条中31号生理小种专化的SCAR检测标记。     

小麦抗条锈病基因定位及分子标记研究进展

本文综述了小麦抗条锈病基因染色体定位及抗条锈病基因分子标记的研究进展,并对几种分子标记技术的应用潜力作了比较分析,特别是对ＳＳＲ、ＩＳＳＲ、ＡＦＬＰ等新型分子标记在小麦遗传育种中的应用前景作了初步探讨。     

温室条件下条形柄锈菌体细胞重组的分子确证

为了获得温室条件下条形柄锈菌发生体细胞重组而导致毒性变异的直接证据,本研究选取7个美国条形柄锈菌小麦专化型菌系和2个美国条形柄锈菌大麦专化型菌系按照夏孢子颜色和专化型与毒性差异组成9对菌系组合,对于室内混合接种产生的子代菌系用具有不同抗性的小麦或大麦品种进行筛选,采用毒性分析及SSR分子标记技术对条形柄锈菌体细胞重组现象进行了研究。对获取的413个单孢子代菌系进行的毒性分析结果显示,有84个单孢子代菌系的毒性谱表现与亲本菌系不同,初步证明体细胞重组过程的存在。SSR标记分析结果显示,11对SSR引物中有6对引物在5对菌系组合的28个毒性谱不同的单孢子代菌系中,检测发现3个单孢菌系的扩增条带与其亲本菌系不同,且表现为亲本菌系扩增条带的重组,为体细胞重组菌系。这一结果从分子水平上证明了条形柄锈菌在室内接种条件下可以通过体细胞重组产生新小种而导致毒性变异。     

条形柄锈菌小麦专化型小种CYR29有性自交及毒性与无毒性基因的遗传分析

由条形柄锈菌小麦专化型(小麦条锈菌)Puccinia striiformisf.sp.tritici引起的条锈病是小麦上重大的生物灾害,严重威胁小麦安全生产.应用抗病品种是防治小麦条锈病最为经济有效的措施,但是条锈菌毒性的频繁变异,常常导致品种抗病性丧失,从而引发条锈病新的大流行.有性生殖是条锈菌毒性变异的重要途径,本...     

小麦条锈菌鉴别寄主抗条锈病基因Yr9的微卫星标记

以含有Yr9的抗条锈病近等基因系Taichung29＊6／Yr9及其轮回亲本Taichung29为材料,用目的基因所在1B染色体上32对微卫星引物对其基因组DNA进行PCR扩增,发现引物Xgwm582在近等基因系与轮回亲本间可扩增出特异性DNA片段。经F2代分离群体177个抗、感单株检测证实,该片段位点与抗条锈病基因Yr9紧密连锁,遗传距离为3．7cM,确定Xgwm582可作为抗条锈病基因Yr9的标记。     

抗条锈病小麦—中间偃麦草异附加系的生化与分子标记

对小麦-中间偃麦草部分双二倍体无芒中4、异附加系C076、宛7107和中国春进行了肽链内切酶（EP-1）等电聚焦电泳。结果表明,肽链内切酶在阳极处有一特异带。肽链内切酶已定位于小麦第7部分同源群,故附加的染色体为第7部分同源群的2条染色体,对中间偃麦草,无芒中4、C076和宛7107进行了RAPD分析。获得了可用于检测C076中外源染色体的3个RAPD标记,即OPI05-800、OPI10-600、OPK01-900。     

小麦秆锈菌生理小种21C3CPH和Ug99 SSR标记的筛选

小麦秆锈病是一种专化性很强的大区远距气传病害,曾造成多个小麦种植国家和地区的毁灭性损失,新的强毒力小种Ug99含有对Sr31等多个重要抗秆锈基因的联合毒性,对我国的小麦生产有巨大潜在威胁,因此,加强小麦秆锈菌生理小种的监测和鉴定是有效防治该病害的基础性研究工作和关键环节。现代分子生物学的迅猛发展,为许多研究提供了新的方法和手段,分子标记技术在区分小麦秆锈菌生理小种方面显示了充分的可行性。本研究利用25对SSR引物对7个小麦秆锈菌主要生理小种进行DNA多态性分析,结果显示,所有特异引物对秆锈菌的扩增结果均呈现出丰富的多态性,秆锈菌的不同生理小种之间存在遗传差异。其中引物SSR180在21C3CPH中扩增出205bp的特异性条带;引物SSR6在Ug99中扩增出170bp的特异性条带,经过多次的重复试验,这些特异性条带均能够比较稳定地重复出现,说明引物SSR180和SSR6可用于小种21C3CPH和Ug99的特异性检测。     

小麦条锈菌分子生态学研究进展

小麦条锈病是危害最严重的小麦流行性病害之一,小麦条锈菌的生态学研究对制定合理的防治策略和抗锈育种具有重要意义.近十几年来,DNA分子标记技术被应用于小麦条锈菌的群体遗传学研究,推动了小麦条锈菌分子生态学研究的快速发展,为揭示小麦条锈菌的群体生态特性开辟了一个新的途径.本文系统介绍小麦条锈菌分子生态学研究的主要进展,并就我国当前研究的局限性和发展趋势进行了分析.     

小麦背景中来自华山新麦草的抗条锈病基因的遗传学分析和分子标记

H9020—17—5是一个通过杂交和回交选育的普通小麦—华山新麦草易位系,接种鉴定表明其对条锈病具有优良抗性。遗传学分析证明易位系H9020—17—5的抗条锈性是由单基因控制的显性性状,抗性基因来自于华山新麦草,暂定名为YrHua。为了标记这个来自华山新麦草的抗条锈病基因,利用H9020—17—5与感病小麦品种铭贤169杂交,建立了F2分离群体。应用81对AFLP引物对119个经条锈菌生理小种CY30接种鉴定的F2单株进行了分析,结果得到两个与YrHua基因连锁的AFLP标记PM14(301)和PM42(249),遗传距离分别为5．4cM和2．7cM,并分别位于目标基因的两侧。将标记片段克隆、测序后,根据序列信息和酶切位点多态性设计特异性引物,将AFLP标记PM14(301)转换成了简单的PCR标记。研究结果为标记辅助育种提供了分子选择工具,同时也为进一步精细定位和图位克隆YrHua基因奠定了基础。     

Quantitative resistance of some Elite wheat lines to Puccinia striiformis f. sp. tritici

Host resistance is the most economical way to manage wheat stripe rust caused by Puccinia striiformis f. sp. tritici. Slow rusting, a type of quantitative resistance, has been reported to last for a long time. Quantitative resistance, in terms of slow rusting parameters including final rust severity (FRS), apparent infection rate (r), relative area under disease progress curve (rAUDPC) and coefficient of infection (CI), was evaluated in a set of 29 wheat genotypes along with susceptible control during 2008–2009 and 2009–2010 cropping seasons. This study was conducted in field plots at Ardabil Agricultural Research Station (Iran) under natural infection conditions with two times artificial inoculation. Artificial inoculation was carried out by yellow rust inoculum having virulent genes against Yr2, Yr6, Yr7, Yr9, Yr22, Yr23, Yr24, Yr25, Yr26, Yr27, YrA and YrSU. Results of mean comparison for resistance parameters showed that lines C-86-1, C-86-2, C-87-1 and C-87-3 along with susceptible had the highest values of FRS, CI, r and rAUDPC, therefore were selected as susceptible lines. The lines C-86-3, C-86-9, C-87-2, C-87-6, C-87-8, C-87-11 and C-87-18 were susceptible at the seedling stage and had low level infection at adult plant stage. Consequently, these lines with low different parameters most probably have slow rusting resistance. The remaining lines had no infection or were at low level of infection. Thus, they were selected as resistant or moderately resistant lines. In this study, correlation coefficient between different parameters of slow rusting was significantly high (r = 0.92–0.99).     

小麦条锈菌冬孢子发生的组织学和超微结构研究

采用冷冻切片技术、光学显微镜和电子显微镜技术,系统研究了小麦条锈菌冬孢子的个体发生过程和超微结构特征。结果表明,小麦条锈菌冬孢子由排列在冬孢子堆基部的双核产孢细胞产生。在发育初期,产孢细胞一端产生突起形成冬孢子芽,随后冬孢子芽经延伸并形成隔膜,依次分化形成冬孢子原基、柄细胞和冬孢子原体。冬孢子原体经有丝分裂后产生隔膜发育形成双核双细胞冬孢子。成熟的冬孢子表面光滑,具有明显加厚的细胞壁,双核融合,原生质密度增加,富含脂肪粒和糖原类物质。在部分冬孢子堆周围还可观察到莲花状包被结构。     

Rates of evolution of avirulence phenotypes and DNA markers in a northwest European population of Puccinia striiformis f. sp. tritici

The effects of evolutionary processes in fungal pathogen populations may occur more rapidly and display larger effects in agricultural systems than in wild ecosystems because of human involvement by plant breeding and crop management. In this study, we analysed the rate of evolution in three lineages of a northwest European population of a biotrophic and asexual reproduced fungal pathogen, Puccinia striiformis f. sp. tritici, causing yellow rust on wheat. Pathogen samples were collected between 1975 and 2002 in the UK and Denmark, and assayed for 14 individual avirulence/virulence alleles and up to 234 amplified fragment length polymorphism (AFLP) primer pairs producing approximately 17,000 AFLP fragments. The large number of fragments and a targeted sampling of isolates allowed a reconstruction of phylogenies in great detail, i.e. no homoplasy and a representation of sequential, evolutionary steps by pathogen samples. A recent, phenotypic loss of avirulence was observed at least once for loci corresponding to P. striiformis f. sp. tritici resistance Yr2, Yr3, Yr4, Yr7, Yr9, and Yr15, whereas Avr6 and Avr17 were lost independently in all three lineages, corresponding to 16 events of loss of avirulence (emergence of virulence). The opposite process, restoration of avirulence, was observed for Yr9 and Yr32. An interpretation of phenotypic changes within lineages as independent mutation events resulted in mutation frequencies from 1.4x10(-6) to 4.1x10(-6) per AFLP fragment (locus) per generation, whereas the effective rate by which a mutation from avirulence to virulence was established in the pathogen population, when subject to selection by host resistance genes, was approximately three orders of magnitude faster.     

Virulence variation of Puccinia striiformis Westend. f. sp. tritici in Pakistan

Yellow rust populations of Pakistan were characterised for their virulence pathotypes/races and pathogenetic variation using seedling evaluation of differential genotypes under glasshouse conditions in Murree (6000 feet above sea level). Differential genotypes comprised a world set, an European set, near isogenic lines and the universally susceptible bread wheat cultivar “Morocco”. Over the two-year study a total of 18 race groups were identified. Out of these 18 race groups, several (68E0, 64E0, 66E0, 70E0, 6E0, 71E0, 6E0, 2E0, 67E0, and 68E16) were found previously. The new race group 70E32 found probably evolved because of mutation from the previously existing 70E16. Virulence frequencies of yellow rust (Yr) resistance genes were also determined on near isogenic lines. The highest virulence frequencies (%) were found for Yr7 (88%), Yr9 (57%), Yr18 (70%), and Yr24 (69%). Virulence frequencies were low for Yr 1 (4%), Yr5 (7%), Yr10 (10%) and Yr15 (4%). Our studies indicated that virulence existed for almost all yr genes, necessitating regular monitoring of the yellow rust populations and intensifying efforts to identify new sources of resistance to this pathogen.     

Early molecular diagnosis and detection of Puccinia striiformis f. sp. tritici in China

Aims: Wheat stripe (yellow) rust, caused by Puccinia striiformis f. sp. tritici (Pst), is the most important foliar disease on wheat in China. Early molecular diagnosis and detection of stripe rust will provide a useful aid to the accurate forecast and seasonal control of this destructive disease. Our objective was to develop PCR assays for the rapid identification and detection of P. striiformis. Methods and Results: The genomic DNA of P. striiformis and P. triticina were amplified by a pair of primers derived from conserved β‐tubulin gene sequence. A 235‐bp specific DNA fragment of P. striiformis was isolated and purified. Based on its sequence, another two primer sets were designed successfully to obtain new sequence‐characterized amplified region (SCAR) markers of P. striiformis, which could be amplified in all test isolates of P. striiformis, whereas no DNA fragment was obtained in other nontarget wheat pathogens. The detection limit of the primer set YR (f)/YR (r1) was 2·20 pg μl^?1. The new SCAR markers of P. striiformis can also be detected in Pst‐infected wheat leaves postinoculated for 2 days. Conclusions: Our assays are significantly faster than the conventional methods used in the identification of P. striiformis. Significance and Impact of the Study: Development of a simple, high‐throughput assay kit for the rapid diagnosis and detection of wheat stripe rust would be anticipated in a further study.     

Phenotypic and Molecular Characterization of Wheat for Slow Rusting Resistance against Puccinia striiformis Westend. f.sp. tritici

Race‐specific resistance of wheat (Triticum aestivum L.) to yellow rust caused by Puccinia striiformis Westend. f.sp. tritici is often short‐lived. Slow‐rusting resistance has been reported to be a more durable type of resistance. A set of sixteen bread wheat varieties along with a susceptible control Morocco was tested during 2004–05 to 2006–07 in field plots at Peshawar (Pakistan) to identify slow rusting genotypes through epidemiological variables including final rust severity (FRS), apparent infection rate (r), area under disease progress curve (AUDPC), average coefficients of infection (ACI) and leaf tip necrosis (LTN). Epidemiological parameters of resistance were significantly (P < 0.01) different for years/varieties in three seasons, while variety × year interactions remained non‐significant. Sequence tagged site (STS) marker, csLV34 analyses revealed that cultivars Faisalabad‐83, Bahawalpur‐95, Suleman‐96, Punjab‐96, Bakhtawar‐93, Faisalabad‐85, Shahkar‐95 and Kohsar‐95 possessed Yr18 linked allele. Faisalabad‐83, Bahawalpur‐95, Suleman‐96, Punjab‐96, Bakhtawar‐93 and Faisalabad‐85 were relatively more stable over 3‐years where FRS, AUDPC and r values reduced by 80, 84 and 70% respectively compared to control Morocco. These six varieties therefore could be exploited for the deployment of Yr18 in breeding for slow rusting in wheat. Both FRS and ACI are suitable parameters for phenotypic selection.     

条形柄锈菌小麦专化型脂肪酶基因PsLIP1的酶学性质分析

脂肪作为条形柄锈菌小麦专化型夏孢子萌发及侵染初期的主要营养物质,酶解产物通过乙醛酸循环转化为糖类成分供病菌生长发育所需。脂肪酶是脂肪代谢的关键酶。基于已测序的基因组序列,利用RT-PCR方法克隆了该病菌脂肪酶基因PsLIP1序列（1 302bp）,编码433个氨基酸的蛋白。在毕赤酵母细胞（GS115）中成功表达PsLIP1后,酶学活性分析显示其最适pH为8.0,最适温度为60℃,此外,发现Zn²⁺、Cu²⁺和Ca²⁺对脂肪酶PsLIP1活力有一定激活作用,Fe²⁺、Mn²⁺则对酶活力有抑制作用。研究结果为进一步解析脂肪酶作用机理奠定基础,同时也为从能量代谢角度制定小麦条锈病防控策略提供理论依据。     

Genetic evidence of local adaptation of wheat yellow rust (Puccinia striiformis f. sp. tritici) within France

Puccinia striiformis f. sp. tritici (PST), a clonal basidiomycete causing yellow rust disease on wheat, has a long record of 'overcoming' cultivar resistance introduced by breeders. Despite the long dispersal capacity of its spores, the French population of PST presents a strong geographical structure, with the presence of a specific pathotype (array of avirulence genes) at high frequencies in the south of France. The genetic diversity underlying this differentiation was analysed by microsatellite and AFLP markers. A total of 213 French isolates belonging to 10 pathotypes collected over a 15-year period were investigated. For each of the 12 microsatellites used, polymorphism resulted from a unique allelic variant associated to the south-specific pathotype. This pathotype was characterized by 40 specific markers over the total of 63 polymorphins detected using 15 AFLP primer combinations. Phylogeographical analysis indicated a strictly clonal structure of the population, and a strong genomic divergence between the northern population and a south-specific clone. Both virulence and molecular data show that the northern French population belongs to the northwestern European population, whereas the southern clone is most likely related to a Mediterranean population, the two subpopulations resulting from the ancient divergence of two clonal lineages. While the virulence complexity in the northern population may be explained by the successive introduction of corresponding resistance genes in cultivars, the maintenance of a simple virulence type in southern France, despite gene flow between the two populations, may be explained in terms of host cultivars repartition and local adaptation to specific host or climatic conditions.