与番茄枯萎病抗病基因Ⅰ-1连锁的AFLP和SSR分子标记

本研究以番茄枯萎病抗病品种‘05045’与感病品种‘051451’为亲本配制杂交组合。用870对AFLP引物及319对SSR引物对348个F2代分离群体进行连锁分析,得到4个与番茄枯萎病抗病基因I-1连锁的AFLP标记和2个SSR标记,分别是E41M60D、E41M62C、E86M36B、E32M44E和SSR108、SSR276,与抗病基因I-1的连锁遗传距离分别为4.7、5.3、8.9、11.5cM和6.1、9.3cM。     

与番茄枯萎病抗病基因I-1连锁的AFLP和SSR分子标记

本研究以番茄枯萎病抗病品种‘05045’与感病品种‘051451’为亲本配制杂交组合。用870对AFLP引物及319对SSR引物对348个F2代分离群体进行连锁分析,得到4个与番茄枯萎病抗病基因I-1连锁的AFLP标记和2个SSR标记,分别是E41M60-D、E41M62-C、E86M36-B、E32M44-E和SSR108、SSR276,与抗病基因I-1的连锁遗传距离分别为4.7、5.3、8.9、11.5cM和6.1、9.3cM。     

番茄抗黄化曲叶病基因Ty-2的分子标记及种质资源初步鉴定

以抗番茄黄化曲叶病毒病材料‘CLN2498D’为父本,感病材料‘早粉2号’为母本,以其F2代为研究对象,采用AFLP及SSR两种标记方法,筛选与Ty-2基因连锁的标记。通过对256对AFLP引物及30对SSR引物的筛选,共获得5个AFLP标记和2个SSR标记与Ty-2基因连锁,其中标记E02M11距离目标基因5.8cM,另有3个标记距离均在10cM以内。将标记E02M11用于30份番茄材料的种质资源筛选,获得12个含有该标记的番茄材料,为番茄抗黄化曲叶病毒育种工作提供基础。     

小麦抗源Sw92抗叶锈病基因遗传及其分子标记

以小麦优异抗源Sw92为父本,感病小麦品种铭贤169为母本,杂交获得F1、F2和BC1代群体。采用我国叶锈菌优势小种PHT对双亲及其杂交世代进行接种鉴定。结果表明,小麦抗源Sw92对叶锈菌小种PHT的抗性系由一对隐性基因所控制。采用简单重复序列(SSR)技术对Sw92携带的抗性基因进行分子标记,共筛选了371对SSR引物,获得2个引物(WMC494、WMC737)可在抗/感池和双亲中扩增出多态性DNA片段。遗传连锁分析结果表明,该抗病基因位于小麦6BS上,与WMC494、WMC737标记的遗传距离分别为3.4cM和15.0cM,不同于6BS上的已知抗叶锈基因Lr36和Lr53,暂命名为LrSw92。     

普通小麦-柔软滨麦草易位系M97抗条锈基因的遗传分析和分子作图

为了明确M97抗条锈性遗传规律,在苗期用7个小麦条锈菌系对M97与感病品种铭贤169的杂交后代F1、F2、F3和BC1代进行抗条锈性遗传分析,并对M97抗Sun11-4的抗条锈基因进行SSR分子标记。M97对Sun11-4和Sun11-11的抗病性均由1对显性基因控制,对CY29、CY30、CY33的抗病性由1显1隐2对基因共同控制,对CY31的抗病性由2对显性基因独立或重叠作用控制。以接种Sun11-4的F2代分离群体构建作图群体,筛选到Xwmc222、Xwmc147、Xbarc229和Xwmc339等4个与抗病基因连锁的SSR标记,其遗传距离分别为3.4、4.8、7.6和12.1 cM。将该抗病基因定位于小麦1DS染色体,且该基因不同于已知的抗条锈基因,暂命名为YrM97。用YrM97两侧遗传距离最近的2个标记Xwmc222和Xwmc147对42个黄淮麦区主栽小麦品种进行分子检测,仅有9.5%的品种具有与YrM97相同的标记位点。     

陆地棉对黄萎病抗性的分子标记研究

 利用陆地棉标准系TM-1和常抗棉2个陆地棉品种杂交并自交,获得109个F₂单株及F_2:3家系为作图群体,以SSR、RAPD和SRAP 3种分子标记进行抗黄萎病性状的分子标记筛选。结果从1611对(条)引物中仅筛选到70对(条)多态性引物,获得75个多态性位点并进行标记间的连锁性分析。75个标记构建了一个包括15个连锁群,全长535 cM的陆地棉品种间分子标记遗传连锁图,标记间平均距离为11.15 cM,有27个标记不能进入任何连锁群。连锁群的标记数最少2个,最多6个;长度从1.0 cM到92.7 cM不等。对其F_2:3家系的成株期抗黄萎病性状即平均病情指数的分布进行分析,显示其呈正态分布,进一步说明陆地棉对黄萎病的抗性为数量遗传;单标记分析及复合区间作图,检测出与抗黄萎病性相关的3个QTL,分别位于第3、5、6连锁群上,贡献率分别为14.15%、3.45%和18.78%。另外,对该群体生长过程中黄萎病不同发病高峰期的病情也进行了分析。     

小麦-柔软滨麦草易位系M853-4抗条锈病基因的分子标记

为揭示小麦-柔软滨麦草易位系M853-4的抗条锈性遗传机制,以易位系M853-4和感病品种铭贤169为亲本制备F2、F3代种子,采用人工接种的方法于温室中接种小麦条锈菌生理小种Su-11,用于测定M853-4及其杂交后代的苗期抗条锈性。结果表明,M853-4对Su-11的抗病性由1对显性和1对隐性基因控制。筛选由1对显性基因控制的F3代分离家系作为SSR标记群体,从320对引物中共找到了4个位于4A染色体上的与该显性基因(暂命名为YrLm2)紧密连锁的微卫星标记Xgwm44、Xwmc650、Barc170和Xwmc718,标记到YrLm2的遗传距离分别为15.0、5.0、3.9和3.1cM,并将YrLm2定位于4A染色体的长臂上,标记结果可用于小麦分子辅助育种。     

大麦抗条纹病基因的定位分析

为发掘大麦中抗条纹病的新基因,采用三明治法通过人工接种大麦条纹病菌Pyrenophora graminea强致病力菌株QWC对甘啤2号(免疫)与Alexis(高感)杂交F_1代及F_2代分离群体进行抗性遗传分析,利用群体分离分析法鉴定与抗病基因连锁的SSR标记,并通过QTL IciMapping软件构建遗传连锁图谱完成对抗病基因的定位。结果显示,甘啤2号与Alexis杂交F_1代对大麦条纹病菌强致病力菌株QWC表现为免疫,F_2代表现3∶1抗感分离,表明甘啤2号对菌株QWC的抗性由1个显性抗性基因控制,将该抗病基因暂命名为Rdg3;该基因位于大麦7H染色体上的SSR标记Bmag206和Bmag7之间,与二者的遗传距离分别为1.78 cM和2.86 cM。经与已定位于7H染色体上的抗病基因比较,发现Rdg3是一个新的抗条纹病基因,可作为大麦抗病育种的新种质资源。     

普通小麦-华山新麦草易位系9020-17-25-6抗条锈性遗传分析及分子标记

为明确普通小麦-华山新麦草易位系9020-17-25-6的抗条锈病基因及其遗传特点,利用中国条锈菌小种CYR29对9020-17-25-6、铭贤169及其杂交后代F1、F2、F3代进行苗期抗条锈性鉴定及遗传分析,选取48条RGAP引物和491对SSR引物对接种CYR29的F2代群体进行筛选,寻找与抗病基因连锁的分子标记。结果表明:9020-17-25-6对CYR29具有良好的抗条锈性,由1对显性基因独立控制,暂定名为Yr Hua9020。筛选到2个RGAP标记(M1和M2)和位于染色体3AS上的4个SSR标记(Xwmc11、Xwmc532、Xcfd79、Xgwm2)与Yr Hua9020连锁,与目的基因的遗传距离分别为6.9、9.5、17.8、12.2、7.2和17.8 c M。与已定位于3A染色体上的抗条锈病基因的比较研究表明,Yr Hua9020是一个与已知基因不同的新的抗条锈病基因。     

小麦抗白粉病基因Pm2的SSR标记筛选

为筛选与小麦抗白粉病基因Pm2紧密连锁的分子标记,将感病品种Chancellor与Pm2的近等基因系杂交,获得F1、F2分离群体,采用分离群体分组法对Pm2进行了微卫星(microsatellite,又称simple sequence repeats,SSR)标记分析.结果表明,定位于小麦5D染色体上的71对SSR引物中有12对引物能在Pm2的近等基因系、Chancellor间稳定地揭示出多态性差异,7对引物Xcfd189、Xcfd29、Xcfd8、Xcfd102、Xcfd7、Xcfd57和Xgwm190分别能在抗病、感病池间和F2分离群体的抗病、感病单株间稳定地扩增出特异性产物.7对引物所扩增的特异谱带分别为:Xcfd189360、Xcfd29190、Xcfd8160、Xcfd102250、Xcfd7200、Xcfd57245和Xgwm190210,它们与Pm2基因间的遗传距离分别为0、1.5、2.3、5.4、10.2、31.5和54.3 cM,其中标记Xcfd189360与Pm2共分离,标记Xcfd29190、Xcfd8160和Xcfd102250与Pm2紧密连锁,可用于Pm2的标记辅助选择.     

小麦-滨麦易位系M8657-1抗条锈病基因遗传分析和分子标记

 M8657-1, one of the wheat translocation lines derived from Leymus mollis Trin. Hara, is possessed of effective resistance at all stages to Su-ll and other dominant races of Puccinia striiformis f. sp. tritici in China. Seedlings of the parents, F₁, and F₂ progeny derived from the cross of M8657-1 (resistant) Mingxian169 (susceptible) were inoculated with Su-ll in greenhouse to identify and map the probable new stripe rust resistance gene. The results suggested that the stripe rust resistance in M8657-1 was conferred by a pair of recessive genes. Simple sequence repeat (SSR) technique was used to detect molecular marker associated with the resistance gene:208 pairs of wheat SSR primers were used to screen the two parents, as well as resistant and susceptible bulks and then three SSR markers were selected for genotyping the F₂ population. The geue, temporarily designated as YrLml, was found to be located on the chromosome 7DL and flanked by three SSR markers GDM67, WMC150 and WMC671, with the genetic distance of 5.0, 9.7 and 11.8cM, respectively.     

小簇麦易位系V9128-3抗条锈病基因的遗传分析和SSR分子标记

 用7个我国当前流行的条锈菌生理小种对V9128-3的抗条锈性进行了评价,表明本易位系对我国优势流行小种具有良好的抗病性。以Su-4对V9128-3与铭贤169配置的F₁、BC₁F₁、F₂及F₃代群体进行了遗传分析,并对其中1个F₂群体进行了SSR标记,再用BC₁F₁群体的部分单株和F₃家系进行连锁标记的初步验证。遗传分析表明了V9128-3对Su-4的抗病性由1对显性核基因独立控制,从219对SSR引物中筛选到2个位于2AL上的该基因YrHV(暂命名)两侧的标记Xgwm356和Xwmc658,遗传距离分别为8.5和5.6cM,所用部分BC₁F₁单株和F₃家系验证了该2个标记与YrHV连锁性。将此标记可用于小麦抗条锈病分子标记辅助育种。     

来自簇毛麦抗条锈病新基因的SSR标记

 用小麦条锈菌条中30号生理小种,对小麦抗病种质小麦-簇毛麦易位系V9128-1和铭贤169的杂交后代进行抗条锈性遗传分析,小麦-簇毛麦易位系V9128-1的抗病性符合1对显性抗条锈病基因控制。并根据F₂抗、感病单株分离比例组建抗感池,用SSR技术寻找与抗病基因连锁的分子标记。从121个SSR引物组合中筛选到2个与抗病基因YrV1(暂命名)紧密连锁的微卫星标记Xgwm566和Xgwm376,遗传距离分别为3.6和5.5cM;因此,该抗条锈病基因位于小麦3B染色体短臂上。这2个标记不仅能在小麦-簇毛麦易位系V9128-1中检测到,而且在抗病基因供体亲本簇毛麦中也能检测到。综合抗病基因来源和分子生物学试验结果,可以推断,YrV1很可能是1个来自簇毛麦并与已知抗条锈病基因不同的新基因。     

Molecular Mapping of the rps1.a Recessive Gene for Resistance to Stripe Rust in BBA 2890 Barley

ABSTRACT Stripe rust, caused by Puccinia striiformis f. sp. hordei, is one of the most important diseases of barley in the south-central and western United States. Growing resistant cultivars is the best approach for controlling the disease. The barley genotype BBA 2890 has all-stage resistance against all races of P. striiformis f. sp. hordei (PSH) identified thus far in the United States. The resistance in BBA 2890 is controlled by a single recessive gene, rps1.a. The objectives of this study were to identify resistance gene analog polymorphism (RGAP) markers for the all-stage resistance gene rps1.a, to map the gene on a barley chromosome using chromosome-specific simple sequence repeat (SSR) markers, and to determine the presence or absence of the flanking RGAP markers for the gene in 24 barley genotypes. Seedlings of the parents and 200 F(8) recombinant inbred lines (RILs) were tested for resistance to pathogen races PSH-14, PSH-48, and PSH-54 in the greenhouse in 2005. Genomic DNA was extracted from the parents and 150 F(8) RILs. The RGAP technique was used to identify molecular markers for the rps1.a gene. Twelve primer pairs generating repeatable polymorphic bands were selected for genotyping the 150 F(8) RILs. A genetic linkage group was constructed for the resistance gene with 13 RGAP markers and four chromosome-specific SSR markers. The four SSR markers mapped the gene on the long arm of barley chromosome 3H. The closest RGAP marker for the resistant allele was within a genetic distance of 2.1 centimorgans (cM). The closest marker for the susceptible allele was 6.8 cM away from the locus. The two closest RGAP markers for the resistant allele detected polymorphisms in 67 and 71% of the 24 barley genotypes when used individually, and detected polymorphism in 88% of the genotypes when used in combination. This information should be useful in incorporating the resistance gene into barley cultivars and in pyramiding the gene with other resistance genes for superior stripe rust resistance.     

普通小麦-华山新麦草易位系H9015-17抗条锈病基因的标记定位

 采用我国当前流行的小麦条锈菌小种和重要致病类型, 在常温条件下对普通小麦-华山新麦草易位系H9015-17进行苗期抗条锈性鉴定, 并用当前主要流行小种CYR32对H9015-17与铭贤169的杂交后代及其双亲进行抗条锈性遗传分析, 以揭示H9015-17抗条锈性遗传基础。结果显示, H9015-17对小麦条锈菌小种CYR31、CYR32、CYR33和致病类型Su11-4、Su11-7、V26、Su11-11均有良好的抗病性, 对当前主要流行小种CYR32的抗病性由1对显性基因控制, 暂命名为YrHua1。 采用分子标记定位技术,筛选到5个与抗病基因YrHua1连锁的RGAP标记(M1、M2、M3、M4和M5)和1个SSR标记(Xgwm292),这些标记与抗病基因YrHua1的遗传距离分别为17.3、15.7、13.1、3.3、2.9和11.2,并将基因YrHua1定位在小麦染色体5DL上。研究结果将为分子标记辅助选择改良小麦抗条锈性提供宝贵的种质材料,建议在抗病育种加以利用。     

Molecular mapping of the crown rust resistance gene rpc1 in barley

ABSTRACT Crown rust of barley, caused by Puccinia coronata var. hordei, occurs sporadically and sometimes may cause yield and quality reductions in the Great Plains region of the United States and Canada. The incompletely dominant resistance allele Rpc1 confers resistance to P. coronata in barley. Two generations, F(2) and F(2:3), developed from a cross between the resistant line Hor2596 (CIho 1243) and the susceptible line Bowman (PI 483237), were used in this study. Bulked segregant analysis combined with random amplified polymorphic DNA (RAPD) primers were used to identify molecular markers linked to Rpc1. DNA genotypes produced by 500 RAPD primers, 200 microsatellites (SSRs), and 71 restriction fragment length polymorphism (RFLP) probes were applied to map Rpc1. Of these, 15 RAPD primers identified polymorphisms between resistant and susceptible bulks, and 62 SSR markers and 32 RFLP markers identified polymorphisms between the resistant and susceptible parents. The polymorphic markers were applied to 97 F(2) individuals and F(2:3) families. These markers identified 112 polymorphisms and were used for primary linkage mapping to Rpc1 using Map Manager QT. Two RFLP and five SSR markers spanning the centromere on chromosome 3H and one RAPD marker (OPO08-700) were linked with Rpc1 and, thus, used to construct a 30-centimorgan (cM) linkage map containing the Rpc1 locus. The genetic distance between Rpc1 and the closest marker, RAPD OPO08-700, was 2.5 cM. The linked markers will be useful for incorporating this crown rust resistance gene into barley breeding lines.     

Identification of allele specific AFLP markers linked with bacterial wilt [Ralstonia solanacearum (Smith) Yabuuchi et al.] resistance in hot peppers (Capsicum annuum L.)

Even though the bacterial wilt is identified as the most destructive disease in hot peppers world-wide, robust molecular markers that facilitate marker assisted selection are absent till date. Kerala Agricultural University (India) has released two hot peppers named Ujwala and Anugraha which show high level field resistance to this pathogen. The variety Anugraha was developed through backcross breeding between a high yielding but highly susceptible variety Pusa Jwala with the highly resistant Ujwala, using Pusa Jwala as a recurrent parent. Thus, Pusa Jwala and Anugraha are near isogenic lines (NILs) differing for the resistance to bacterial wilt only and the resistance is governed by a homozygous recessive (rr) gene action. The F₁s of Anugraha × Pusa Jwala were selfed to generate the segregating F₂ population. The F₂ population has been field screened, 10 highly susceptible and 10 most resistant plants were identified and DNA from these plants were bulked separately. Bulked segregant analysis with AFLP primer combination EcoACT + MseCAC was done using the DNA from donor parent Ujwala, susceptible parent Pusa Jwala, resistant parent Anugraha, bulked susceptible F₂ and bulked resistant F₂ plants. On resolution using capillary electrophoresis system in genetic analyzer, the AFLP products have yielded three polymorphic bands (103, 117, and 161 bp) which were linked with the resistant recessive allele and three polymorphic bands (183, 296, 319 bp) linked with the dominant susceptible allele of the bacterial wilt resistance gene. The results were confirmed through co-segregation analysis in most resistant and susceptible plants of F₂ segregating population.     

Molecular mapping of the leaf rust resistance gene rph6 in barley and its linkage relationships with rph5 and rph7

ABSTRACT The barley cv. Bolivia carries two leaf rust (Puccinia hordei) resistance genes, Rph2 and Rph6, and is the only known source of the latter gene. A resistant line (Bolivia-Rph6) carrying Rph6 only was obtained in the F(4) generation of a cross between cv. Bolivia and the susceptible cv. Bowman via progeny testing with differential isolates of the leaf rust pathogen. Genetic analyses and bulk segregant analysis using amplified fragment length polymorphism (AFLP) and restriction fragment length polymorphism (RFLP) markers localized Rph6 on the short arm of barley chromosome 3H at a distance of 4.4 centimorgans (cM) distal from RFLP marker MWG2021 and 1.2 cM proximal from RFLP marker BCD907. The allelic relationship of Rph6 to other leaf rust resistance genes mapping to this region of chromosome 3H (namely Rph5 and Rph7) were tested using crosses among cvs. Magnif 102 (carrying Rph5), Bolivia-Rph6 (Rph6), and Cebada Capa (Rph7). Segregation analyses indicated that Rph6 is allelic to Rph5 and closely linked to Rph7. The data generated from this study will facilitate breeding for leaf rust resistance via marker-assisted selection and provide a starting point for positional gene cloning.     

Identification and Molecular Mapping of a Gene in Wheat Conferring Resistance to Mycosphaerella graminicola

ABSTRACT Septoria tritici leaf blotch (STB), caused by the ascomycete Mycosphaerella graminicola (anamorph Septoria tritici), is an economically important disease of wheat. Breeding for resistance to STB is the most effective means to control this disease and can be facilitated through the use of molecular markers. However, molecular markers linked to most genes for resistance to STB are not yet available. This study was conducted to test for resistance in the parents of a standard wheat mapping population and to map any resistance genes identified. The population consisted of 130 F(10) recombinant-inbred lines (RILs) from a cross between the synthetic hexaploid wheat W7984 and cv. Opata 85. Genetic analysis indicated that a single major gene controls resistance to M. graminicola in this population. This putative resistance gene is now designated Stb8 and was mapped with respect to amplified fragment length polymorphism (AFLP) and microsatellite markers. An AFLP marker, EcoRI-ACG/MseI-CAG5, was linked in repulsion with the resistance gene at a distance of approximately 5.3 centimorgans (cM). Two flanking microsatellite markers, Xgwm146 and Xgwm577, were linked to the Stb8 gene on the long arm of wheat chromosome 7B at distances of 3.5 and 5.3 cM, respectively. The microsatellite markers identified in this study have potential for use in marker-assisted selection in breeding programs and for pyramiding of Stb8 with other genes for resistance to M. graminicola in wheat.     

黄瓜枯萎病菌遗传多样性的AFLP分析

黄瓜枯萎病是由半知菌亚门尖孢镰刀菌黄瓜专化型(Fusarium oxysporumf.sp.cucumainum Owen)侵染引起的一种土传病害,是影响黄瓜生产的最主要病害之一[1].近年来随着分子生物学技术的迅速发展,国内外学者对于病原真菌的遗传多样性做了大量的研究,Wang等[2]对影响黄瓜枯萎病菌AFLP技术体系的多种因素作了探讨,得到了1种适合于黄瓜枯萎病菌AFLP分析的优化体系;Duan等[3]应用RAPD、ISSR和AFLP标记揭示出了西瓜枯萎病菌株在分子水平上的遗传多样性.