条锈菌诱导的小麦病程相关蛋白TaPR10基因的克隆及特征分析

 【目的】克隆条锈菌诱导的小麦病程相关蛋白PR10基因,研究其在小麦成株抗条锈病防御反应中的作用。【方法】采用电子克隆、RT-PCR技术,从条锈菌侵染的小麦兴资9104中,分离病程相关蛋白10基因;采用生物信息学技术预测分析该基因的DNA序列结构及其编码蛋白的保守域及基本特性;利用实时荧光定量RT-PCR技术,分析该基因在小麦成株期和苗期受条锈菌CYR32侵染后的表达情况。【结果】分离到病程相关蛋白10基因,命名为TaPR10,ORF长483 bp,编码由160个氨基酸组成的蛋白质TaPR10;TaPR10不含跨膜区、无信号肽、定位在胞内,除具有典型的病程相关蛋白Bet_v_I家族保守结构域外,还有其它4类功能保守域;与小麦、高粱、玉米和水稻等4种植物PR10蛋白的氨基酸序列相似性在80%左右;TaPR10 DNA序列内部存在188至271位84 bp的内含子序列,其拼接位点序列具有GT-AG双核苷酸序列;TaPR10基因表达分析的结果表明,TaPR10基因在成株期和苗期反应中表达量均上调,成株期表达高于苗期。【结论】首次分离到一个条锈菌CYR32诱导的小麦TaPR10基因,该基因可能参与了小麦成株抗条锈病防御反应。     

Pathogenesis-related protein genes involved in race-specific all-stage resistance and non-race specific high-temperature adult-plant resistance to Puccinia striiformis f. sp. tritici in wheat

Interactions of the stripe rust pathogen (Puccinia striiformis f. sp. tritici) with wheat plants activate a wide range of host responses. Among various genes involved in the plant-pathogen interactions, the expressions of particular pathogenesis-related (PR) protein genes determine different defense responses. Different types of resistance have been recognized and utilized for developing wheat cultivars for resistance to stripe rust. All-stage resistance can be detected in seedling stage and remains at high levels throughout the plant growth stages. This type of resistance is race-specific and not durable. In contrast, plants with only high-temperature adult-plant (HTAP) resistance are susceptible in seedling stage, but become resistant when plants grow older and the weather becomes warmer. HTAP resistance controlled by a single gene is partial, but usually non-race specific and durable. The objective of this study was to analyze the expression of PR protein genes involved in different types of wheat resistance to stripe rust. The expression levels of 8 PR protein genes (PR1, PR1.2, PR2, PR3, PR4, PR5, PR9 and PR10) were quantitatively evaluated at 0, 1, 2, 7 and 14 days after inoculation in single resistance gene lines of wheat with all-stage resistance genes YrTr1, Yr76, YrSP and YrExp2 and lines carrying HTAP resistance genes Yr52, Yr59, Yr62 and Yr7B. Races PSTv-4 and PSTv-37 for compatible and incompatible interactions were used in evaluation of PR protein gene expression in wheat lines carrying all-stage resistance genes in the seedling-stage experiment while PSTv-37 was used in the HTAP experiment. Analysis of quantitative real-time polymerase chain reaction (qRT-PCR) revealed that all of the PR protein genes were involved in the different types of resistance controlled by different Yr genes. However, these genes were upregulated at different time points and at different levels during the infection process among the wheat lines with different Yr genes for either all-stage resistance or HTAP resistance. Some of the genes were also induced in compatible interactions, but the levels were almost always higher in the incompatible interaction than in the compatible interaction at the same time point for each Yr gene. These results indicate that both allsalicylic acid and jasmonate signaling pathways are involved in both race-specific all-stage resistance and non-race specific HTAP resistance. Although expressing at different stages of infection and at different levels, these PR protein genes work in concert for contribution to different types of resistance controlled by different Yr genes.     

小麦条锈病和白粉病成株抗性研究进展与展望

 利用成株抗性是小麦抗病育种的重要方向,本文综述了小麦条锈病和白粉病成株抗性鉴定方法、基因定位和克隆及其在育种中的应用。将报道的72个条锈病成株抗性数量性状遗传位点（quantitative trait loci,QTL）和82个白粉病成株抗性QTL整合到一张连锁图谱上,控制2种病害的基因簇（≥5个QTL）有8个,其中位于7DS的Yr18/Lr34/Pm38和1BS的Yr29/Lr46/Pm39对条锈病、叶锈病和白粉病均表现成株抗性,位于4DL的Yr46/Lr67位点可能也对白粉病表现成株抗性,Yr18/Lr34/Pm38和Yr36已被克隆,Yr29/Lr46/Pm39的克隆已取得良好进展,为培育兼抗和成株抗性相结合的品种提供了可用基因。总结了成株抗性在中国小麦育种中的应用现状,并用实例证实了培育成株抗性品种的可行性,建议对兼抗条锈病和白粉病成株抗性的咸农4号和小偃6号等进行遗传分析,育种工作者和品种审定部门需要转变观念,将成株抗性利用作为国内条锈病和白粉病抗性育种的重要内容。     

CIMMYT小麦PBW343和Muu中条锈和叶锈成株抗性QTL分析

为发掘CIMMYT小麦品种PBW343和Muu中条锈和叶锈成株抗性基因,以PBW343与Muu杂交的146个F6代重组自交系为材料,种植于田间调查鉴定,并利用31个SSR标记、16个EST标记和502个DArT(Di-versity Arrays Technology)标记构建连锁图,采用复合区间作图法进行小麦条锈病和叶锈病的成株抗性QTL分析,发现了2个控制小麦抗条锈病和1个控制小麦抗叶锈病的QTL,分别位于2AL、2BL和5BL上,解释8.89%,10.81%和12.82%的表型变异,3个QTL均来自小麦品种Muu。这2个小麦抗条锈QTL和1个抗叶锈QTL的发掘,将为小麦抗病育种提供理论和技术支持。     

四川地方小麦品种条锈病抗性研究

【目的】利用获得的小麦条锈病成株抗性"一致性"QTL位点SSR分子标记,结合田间抗性表型鉴定,揭示四川地方小麦品种的条锈病抗性遗传多样性。【方法】通过QTL元分析技术,将已定位的小麦条锈病成株抗性QTL整合于同一张连锁图谱,获得条锈抗性"真实"QTL区段。并利用QTL区段内的SSR标记,结合田间条锈病抗性表型鉴定,对64份四川地方小麦品种抗条锈病多样性进行研究。【结果】通过元分析,获得7个控制小麦条锈病成株抗性MQTL。成株期条锈病抗性鉴定发现11份四川地方小麦品种表现为免疫或近免疫,其发病严重度、平均普遍率和病情指数低于10%。标记/性状关联分析发现3个与四川小麦地方品种条锈病成株抗性存在显著关联的SSR标记位点。【结论】四川地方小麦品种条锈病抗性表现差异显著;小麦条锈病成株抗性关联SSR标记的获得为利用条锈病抗性基因提供了分子标记辅助选择手段。     

QTLMapping forAdult-Plant Resistance to Stripe Rust in a CommonWheat RIL Population Derived from Chuanmai 32/Chuanyu 12

Stripe rust, caused by Puccinia striiformis f. sp. tritici, is a devastating wheat disease worldwide. The Chinese wheat cultivar Chuanmai 32 has shown stable resistance to stripe rust for 10 yr in Sichuan Province, a hotspot for stripe rust epidemics. The objective of the present study was to map quantitative trait loci (QTL) for adult-plant resistance (APR) to stripe rust in a population of 140 recombinant inbred lines (RILs) derived from Chuanmai 32/Chuanyu 12. Field trials were conducted in Chengdu and Yaan, Sichuan, from 2005 to 2008, providing stripe rust reaction data for 6 environments. 797 simple sequence repeat (SSR) markers were screened for association with stripe rust reaction, initially through bulked segregant analysis (BSA). Based on the mean disease values averaged across environments, the broad-sense heritability of maximum disease severity (MDS) was 0.75. Two QTLs for stripe rust resistance were detected by composite interval mapping (CIM). They were designated QYr.caas-3BL and QYr.caas-3BS and explained from 6.6 to 20.1%, respectively, of the phenotypic variance across environments. QYr.caas-3BL came from Chuanmai 32; QYr.caas-3BS with lower effect was from the susceptible parent Chuanyu 12. Both QTLs appear to be new.     

Identification of QTL for adult plant resistance to stripe rust in bread wheat line C33

Stripe rust, caused by Puccinia striiformis f. sp. tritici, is a serious disease in bread wheat(Triticum aestivum L.). Identification and use of adult plant resistance(APR) resources are important for stripe rust resistance breeding. Bread wheat line C33 is an exotic germplasm that has shown stable APR to stripe rust for more than 10 years in Sichuan Province of China. Here, 183 recombinant inbred lines(RILs) derived from the cross between C33 and a susceptible line X440 were genotyped with diversity arrays technology(DArT) markers to identify resistance quantitative trait locus(QTL). Field trials were conducted in five years at Chengdu and Xindu of Sichuan Province, using maximum disease severity(MDS) as stripe rust reaction phenotypes. A total of four quantitative trait loci(QTLs) were detected, respectively designed as QYr.saas-3 AS, QYr.saas-5 AL, QYr.saas-5 BL, and QYr.saas-7 DS, explaining 4.14–15.21% of the phenotypic variances. QYr.saas-5 BL and QYr.saas-7 DS were contributed by C33. However, the level for stripe rust resistance contributed by them was not strong as C33, suggesting the presence of other unidentified QTLs in C33. QYr.saas-7 DS corresponded to Yr18 and QYr.saas-5 BL remains to be formally named. The RIL lines carrying combinations QYr.saas-5 AL, QYr.saas-5 BL, and QYr.saas-7 DS showed comparability resistance with C33. The present study provides resources to pyramid diverse genes into locally adapted elite germplasm to improve the stripe rust resistance of bread wheat.     

80份国外春小麦种质资源抗条锈性评价

【目的】小麦条锈病是由小麦条锈菌（Puccinia striiformis f. sp. tritici,Pst）引起的世界范围内小麦重要病害之一,培育和种植抗病品种是控制该病害的最有效策略。评价80份国外春小麦种质资源对中国当前小麦条锈菌流行小种的抗条锈性,为中国小麦抗条锈病育种提供依据和抗源。【方法】应用中国流行小麦条锈菌生理小种CYR29、CYR31、CYR32、CYR33以及致病类型PST-HY8和PST-V26对80份国外小麦种质资源进行苗期温室抗病性鉴定,以铭贤169和AvS为感病对照品种;并于2013年和2014年分别在陕西省杨凌和甘肃省天水进行田间成株期抗病性鉴定。根据苗期和田间成株期的抗病性鉴定结果对其进行抗病类型分类和评价。【结果】80份小麦种质资源的抗病类型可分为3类。第1类为全生育期抗病类型,有8份。其中PI660067、PI660119和PI660122在苗期和田间成株期均表现较高水平的抗病性。其余5个品系PI660056、PI607839、PI591045、TA5602和PI660064在苗期则对个别小种表现感病,并且在不同年份和不同测试地点成株期也表现感病。第2类为成株抗病类型,有28份。其苗期对所有测试小种均表现感病,有23份在田间成株期均表现抗病。但PI660075、PI660083、PI660085、PI660097和PI660107在不同年份和不同测试地点成株期表现感病。第3类为兼具成株期和对部分中国小种失去抗性的全生育期抗病类型,有44份,其苗期至少对一个测试小种表现抗病。有37份在田间成株期均表现抗病。但PI660065、PI660076、PI660079、PI660080、PI660095、PI660096和PI610750在不同年份和不同测试地点成株期表现感病。【结论】80份国外小麦种质资源中大部分对中国小麦条锈菌流行小种表现优良的抗病性。这些种质资源可作为抗源在今后抗病育种中加以利用,将丰富中国小麦抗条锈病基因的多样性。可能由于不同年份田间流行小种不同,造成一些成株抗病品系在不同年份和不同测试地点表现感病,由此推测成株抗病性可能也具有小种专化性。     

Study of the Adult-Plant Resistance at Different Growth Stages to Stripe Rust in Wheat

Components of resistance were investigated for five adult-plant resistant (APR) wheat (Triticum aestivum) cultivars at six growth stages and two temperatures in the greenhouse, and disease progress in a field trial. Chinese cultivar Chuanyu 12displayed high to intermediate infection type (IT) in the greenhouse but was highly resistant in the field. Weebill showed an intermediate IT in the greenhouse and also in the field. Chpaio, Tukuru and Saar, known to carry combinations of Yr18and 2-3 additional minor genes, were highly resistant in both experiments. Greenhouse experiments indicated that the lower IT of APR cultivars initiated at tillering stage. Latent periods (LP) for APR cultivars were generally longer as the growth stage progressed. We conclude that APR to stripe rust can be best characterized in field trials although significant correlations are seen between field severity and IT and LP measured in the greenhouse.     

小麦抗条锈病基因对中国条锈菌主要流行小种的抗性分析

[目的]培育和广泛应用抗病小麦品种是防治条锈病最经济有效和环境友好的措施.由于条锈菌(Puccinia striiformis f.sp.tritici,Pst)群体中毒性变异频繁,发生新生理小种常导致主栽品种抗病性'丧失',引发条锈病大规模流行,严重威胁我国主粮安全供给.本研究通过监测和评价已知抗条锈病基因对我国目前...     

小麦品种成株期抗条锈性表达生育期的研究

 对5个携带不同成株期抗性的小麦品种在温室和田间成株期抗病性的研究表明，Chuanyu 12在温室条件下表现感病到中度感病，而在田间表现抗病；携带慢锈性抗性的品种Weebill在温室和田间都表现中度抗病；携带Yr18和2~3个微效抗性基因的品种Chpaio, Tukuru 和 Saar，在温室和田间都表现高度抗病。在温室条件下，对在不同生育期5个抗病品种的反应型和潜育期的研究表明，反应型在分蘖期开始降低，而潜育期随着生育期的发展表现延长。虽然田间严重度和温室反应型、潜育期之间有良好的相关性，成株期抗性的鉴定仍最好在田间进行。     

Genetic analysis of adult plant,quantitative resistance to stripe rust in wheat landrace Wudubaijian in multi-environment trials

Stripe rust, caused by Puccinia striiformis f. sp. tritici(Pst), is one of the most destructive diseases on wheat worldwide.Wudubaijian, a wheat landrace released from Gansu Province in China since 1950, exhibits adult-plant resistance to stripe rust for several decades. To elucidate the genetic basis of stripe rust resistance, Wudubaijian was crossed with the high susceptible cultivar Mingxian 169, and stripe rust tests of both parents and the F2:3 lines were conducted in four environments of Y...     

219个小麦品种成株期慢锈性品种的筛选

[目的]从几大小麦产区主栽品种中筛选出慢锈品种。[方法]用当前条锈菌优势小种接种孕穗期的小麦,计算各品种的潜育期、病情指数及千粒重损失率,从而推算出各品种的慢锈类型。[结果]219个参试品种中筛选出5个免疫品种、67个高度慢锈品种、66个中度慢锈品种、11个耐锈品种和70个感病品种。[结论]在主栽品种中感病类型占有相当的比例,在今后的育种工作中,应多纳入持久抗病种质,逐步淘汰感病种质。     

成株抗条锈性小麦品种的筛选及应用前景

通过三年大田及室内试验,筛选出了我国成株抗条锈性的小麦品种47个.这些品种的抗性可分为三种类群:(1)高抗至免疫类型;(2)中抗类型;(3)介于高抗至中抗之间的类型,该类型的抗性表现取决于环境条件.并对成株抗条锈性品种的应用前景进行了评价.     

152份黄淮海麦区小麦农家品种抗条锈性评价及重要条锈病抗性基因的分子检测

【目的】黄淮海麦区是中国最重要的小麦生产基地和条锈病流行区,从该麦区小麦农家品种中鉴定一批对当前小麦生产上流行的条锈病生理小种具有稳定抗性的优良种质,并了解其携带当前抗病育种利用的重要条锈病抗性基因分布,为进一步在条锈病抗性育种中有效利用和发掘其抗性基因提供材料基础。【方法】应用当前中国小麦生产上毒性强、流行频率高的条锈菌生理小种条中32(CYR32)和条中34(CYR34)对152份来源于黄淮海麦区小麦农家品种进行苗期抗性鉴定,并于2016年和2018年分别在四川崇州和绵阳利用由条锈菌CYR32、CYR33、CYR34、水源11-4、水源11-5组成的混合小种进行人工接种和成株期抗性鉴定;同时利用Yr9、Yr10、Yr18、Yr24/26、Yr30、Yr36、Yr39、Yr41、Yr48、Yr65、Yr67、Yr80和Yr81共13个小麦育种和生产上应用的重要已知条锈病抗性基因的标记进行分子检测,推测其可能携带的抗性基因。【结果】9份农家品种对CYR32具有苗期抗性,9份对CYR34具有苗期抗性;其中,2份种质在苗期对CYR32和CYR34均表现抗性。35份农家品种表现为稳定的成株期...     

中国40个小麦农家品种和甘肃南部40个生产品种抗条锈病基因推导

【目的】明确中国小麦农家品种与甘肃南部生产品种抗条锈性类型及可能含有的抗性基因和遗传多样性,为抗源的选择与利用提供参考。【方法】苗期于自控温室内使用中国当前小麦条锈菌（Puccinia striiformis f. sp. tritici）流行小种CYR32,成株期于大田病圃使用当前主要流行小种和重要致病类型共12个菌系组成的混合菌种对80个供试材料进行抗病性鉴定与评价。基因推导在温室用25个毒性谱不同的小麦条锈菌系于苗期接种30个已知抗条锈病基因载体品系及对照铭贤169、17个国际鉴别寄主和供试品种,根据供试品种和标准品系对不同菌系的侵染型,对农家品种和生产品种进行抗性谱比对分析和系谱分析,解析其可能含有的抗条锈病基因或基因组合及抗性谱,并通过NTSYSpc-2.2软件计算遗传相似系数,以UPGMA法进行聚类分析,将其抗性归类。【结果】供试品种大多具有良好的成株期抗性,除清农1号、清农2号、红壳小麦(2)在成株期表现感病,兰天3号、兰天4号、兰天6号等20个品种在成株期表现慢锈性外,其他品种均表现中抗至免疫的抗性水平。品种抗性多由全生育期抗性、部分由成株抗性提供。甘肃生产品种中抗病基因以Yr9、Yr24、Yr26为主,有的还含有其他未知抗条锈病基因。其中,兰天1号、兰天14号、兰天17号、兰天21号、清农4号等含Yr9;兰天24号、中梁04335、天选51号可能含Yr24;兰天17号、兰天23号、兰天25号、中梁17号、中梁04260、天选43号、天选48号可能含Yr26。农家品种中有19份材料含有未知抗条锈病基因,其余21份材料不能确认含已知抗病基因Yr1、Yr2、Yr4、Yr6、Yr7、Yr8、Yr40,可能含有未知基因。聚类分析发现,甘肃生产品种大都抗性谱较宽,遗传相似性较高;40份材料的抗条锈性相似系数范围在0.30-1.00,在抗性相似系数为0.70水平上可分为3大类,其中兰天15号单独聚为1类,清农1号和清农2号聚为1类,其余37份材料聚为1大类。农家品种抗性谱宽窄不一,显示出遗传多样性水平较高;40份材料的抗条锈性相似系数范围在0.38-1.00,在抗性相似系数为0.70水平上可分9大类。【结论】供试品种均有良好的抗条锈性,相对于甘肃南部生产品种,供试的农家品种更具丰富的遗传多样性和有效抗条锈病基因,可作为抗源在育种中加以利用。     

Genome-wide association analysis for stripe rust resistance in spring wheat(Triticum aestivum L.) germplasm

Stripe rust is a continuous threat to wheat crop all over the world. It causes considerable yield losses in wheat crop every year. Continuous deployment of adult plant resistance(APR) genes in newly developing wheat cultivars is the most judicious strategy to combat this disease. Herein, we dissected the genetics underpinning stripe rust resistance in Pakistani wheat germplasm. An association panel of 94 spring wheat genotypes was phenotyped for two years to score the infestation of stripe rust on each accession and was scanned with 203 polymorphic SSRs. Based on D' measure, linkage disequilibrium(LD) exhibited between loci distant up to 45 c M. Marker-trait associations(MTAs) were determined using mixed linear model(MLM). Total 31 quantitative trait loci(QTLs) were observed on all 21 wheat chromosomes. Twelve QTLs were newly discovered as well as 19 QTLs and 35 previously reported Yr genes were validated in Pakistani wheat germplasm. The major QTLs were QYr.uaf.2 AL and QYr.uaf.3 BS(PVE, 11.9%). Dissection of genes from the newly observed QTLs can provide new APR genes to improve genetic resources for APR resistance in wheat crop.     

条锈菌诱导的小麦TaHin1的克隆与表达特征分析

【目的】克隆受条锈菌诱导的小麦Hin1,研究其在小麦抗条锈病防御反应以及抗非生物胁迫过程中的作用。【方法】通过电子克隆、RT-PCR方法,从条锈菌侵染的小麦水源11中分离出一个编码HIN1的cDNA序列;通过生物信息学工具对DNA序列及其编码的蛋白序列进行分析;利用实时荧光定量PCR分析该基因在小麦不同器官、与条锈菌互作、外源激素处理以及非生物胁迫下的表达情况。【结果】分离得到小麦Hin1,命名为TaHin1,开放阅读框为642bp,编码213个氨基酸,分子量为23.24kD,等电点8.67,具有信号肽和HIN1结构域,可能为分泌蛋白;与高粱和水稻HIN1同源性达80%;表达分析结果表明,TaHin1在小麦根、茎、叶组织中表达差异不显著,在小麦与条锈菌互作时,仅在非亲和组合中被诱导表达;外源植物激素水杨酸、茉莉酸诱导TaHin1上调表达,脱落酸诱导其下调表达,而乙烯不诱导其表达;高盐、干旱、机械损伤以及低温等非生物胁迫下,TaHin1均上调表达。【结论】首次克隆到一个受条锈菌诱导的小麦TaHin1,可能通过水杨酸、茉莉酸信号途径参与了小麦对条锈菌的防御反应,而脱落酸在防御反应中起负调控作用,该基因在小麦对非生物胁迫的抗逆应答过程中也发挥着一定作用。     

Seedling and adult plant resistance to leaf rust in 46 Chinese bread wheat landraces and 39 wheat lines with known Lr genes

Wheat leaf rust,caused by Puccinia triticina(Pt),is an important foliar disease that has an important influence on wheat yield.The most economic,safe and effective way to control the disease is growing resistant cultivars.In the present study,a total of 46 wheat landraces and 34 wheat lines with known Lr(leaf rust resistance)genes were inoculated with 16Pt pathotypes for postulating seedling resistance gene(s)in the greenhouse.These cultivars and five wheat differential lines with adult plant resistance(APR)genes(Lr12,Lr22b,Lr34,Lr35 and Lr37)were also evaluated for identification of slow rusting resistance in the field trials in Baoding,Hebei Province of China in the 2014–2015 and 2015–2016 cropping seasons.Furthermore,10 functional molecular markers closely linked to 10 known Lr genes were used to detect all the wheat genotypes.Results showed that most of the landraces were susceptible to most of the Pt pathotypes at seedling stage.Nonetheless,Lr1 was detected only in Hongtangliangmai.The field experimental test of the two environments showed that 38 landraces showed slow rusting resistance.Seven cultivars possessed Lr34 but none of the landraces contained Lr37 and Lr46.Lr genes namely,Lr9,Lr19,Lr24,Lr28,Lr29,Lr47,Lr51 and Lr53 were effective at the whole plant stage.Lr18,Lr36 and Lr45 had lost resistance to part of pathotypes at the seedling stage but showed high resistance at the adult plant stage.Lr34 as a slowing rusting gene showed good resistance in the field.Four race-specific APR genes Lr12,Lr13,Lr35 and Lr37 conferred good resistance in the field experiments.Seven race-specific genes,Lr2b,Lr2c,Lr11,Lr16,Lr26,Lr33 and LrB had lost resistance.The 38 landraces showed slow rusting resistance to wheat leaf rust can be used as resistance resources for wheat resistance breeding in China.