Molecular mapping and identification of QTL's associated to oat crown rust partial resistance

Summary Molecular mapping is a promising strategy for studying and understanding traits with complex genetic control, such as partial resistance to oat crown rust. The objectives of this research were to develop molecular maps from the progenies of the cross UFRGS7 (susceptible) × UFRGS910906 (partially resistant) and to identify QTLs (quantitative trait loci) associated to partial resistance to oat crown rust in two generations of that population.DNA of 86 genotypes of the F₂ and 90 genotypes of the F₆ UFRGS7 × UFRGS910906 population were used to generate AFLP markers. Molecular maps were constructed using Mapmaker Exp. 3.0 and QTLs for partial resistance to oat crown rust were identified with Mapmaker/QTL software. Five hundred and fifty seven markers in the F₂ and 243 markers in the F₆ generations were identified. The F₂ map integrated 250 markers in 37 linkage groups. The F₆ map integrated 86 markers in 17 linkage groups.Five QTLs were identified for partial resistance to oat crown rust in the F₂ generation and three QTLs in the F₆. The QTL identified on F₆ through the PaaaMctt340 AFLP marker showed consistency across two environments and two generations (F₄ and F₆), and appear to have potential for marker-assisted selection in oat.     

利用3个F_2群体整合高密度栽培种花生遗传连锁图

遗传图谱的构建及整合是开展花生分子育种研究的基础,利用多个作图群体整合遗传图谱是解决图谱标记密度低的有效途径。本研究采用基于锚定SSR标记的作图策略,构建3个F_2群体3张遗传连锁图,利用Join Map 3.0软件整合图谱,获得一张包含20个连锁群、792个位点、总遗传距离为2079.50 c M,标记间平均距离为2.63 c M的整合图谱,各连锁群标记数在20~66个之间,遗传距离在59.10~175.80 c M之间。将3个分离群体中检测到的与荚果及种子大小相关的QTL区段与整合连锁图的标记比较发现,各群体中检测到的位于各染色体上的QTL在整合图谱中都能出现,有些QTL标记区间在整合图谱中存在更多的标记,为今后利用这些标记进行精细定位奠定了基础。     

Clustering of amplified fragment length polymorphism markers in a linkage map of rye

Amplified fragment length polymorphisms (AFLPs) are now widely used in DNA fingerprinting and genetic diversity studies, the construction of dense genetic maps and in fine mapping of agronomically important traits. The AFLP markers have been chosen as a source to extend and saturate a linkage map of rye, which has previously been generated by means of restriction fragment length polymorphism, random amplified polymorphic DNA, simple sequence repeat and isozyme markers. Gaps between linkage groups, which were known to be part of chromosome 2R, have been closed, thus allowing the determination of their correct order. Eighteen EcoRI‐MseI primer combinations were screened for polymorphism and yielded 148 polymorphic bands out of a total of 1180. The level of polymorphism among the different primer combinations varied from 5.7% to 33.3%. Eight primer combinations, which revealed most polymorphisms, were further analysed in all individuals of the F₂ mapping population. Seventy‐one out of 80 polymorphic loci could be integrated into the linkage map, thereby increasing the total number of markers to 182. However, 46% of the mapped AFLP markers constituted four major clusters located on chromosomes 2R, 5R and 7R, predominantly in proximity to the centromere. The integration of AFLP markers caused an increase of 215 cM, which resulted in a total map length of almost 1100 cM.     

Location of genes for common bunt resistance in the European winter wheat cv. Trintella

The location of new genes for resistance to common bunt in wheat is valuable for gene pyramiding in breeding. For this purpose, the genetics of the relatively high level of resistance in the European winter wheat variety Trintella was investigated using a doubled haploid mapping population of a cross between Trintella and the susceptible variety Piko. The population was scored for bunt infection in the field for 2 years following inoculation with a mixture of teliospores of Tilletia tritici and T. laevis. A genetic map consisting of 29 linkage groups was constructed using polymorphic simple sequence repeat markers. This map was used for QTL analysis, and in both years, results indicated that resistance to common bunt could mostly be attributed to a gene on chromosome 1B, near to the centromere and closest to marker Xgwm273 on the short arm. Additionally, in 2008, smaller QTL effects were ascribed to chromosomes 7A and 7B, and another smaller QTL effect to chromosome 5B in 2009 only.     

RFLP/AFLP mapping of a brown planthopper (Nilaparvata lugens Stål) resistance gene Bph1 in rice

We have constructed a linkage map of the rice brown planthopper (BPH)resistance gene, Bph1. RFLP and AFLP markers were selected by thebulked segregant analysis and used in the mapping study of 262 F₂sthat were derived from a cross of `Tsukushibare', a susceptible japonica cultivar, and `Norin-PL3', an authentic japonicaBph1-introgression line. Twenty markers were mapped within a 28.9-cMregion containing the Bph1 locus on the long arm of rice chromosome12. Combining the result of segregation analysis of BPH resistance by themass seedling test and that of the markers, the Bph1 locus wasmapped within a 5.8-cM region between two flanking markers. The closestAFLP markers, em5814N and em2802N, was at 2.7 cM proximal to theBph1 locus. Together with the previously constructed high-resolutionmap of bph2 locating the locus at ca. 10 cM proximal to the Bph1 locus, this improved version of the linkage map would facilitatepyramiding these two important BPH resistance genes.     

The first genetic maps of cashew (Anacardium occidentale L.)

Cashew (Anacardium occidentale) is a widespread tropical tree crop that is grown primarily for its nuts and has a global production of over 2 million Mt. In spite of its economic importance to many countries, however, no linkage map containing STS anchor sites has yet been produced for this species. This is largely attributable to a prolonged juvenile phase of the tree (limiting mapping to F₁ progenies) and difficulty in effecting sufficient hand-pollinations to create mapping populations of effective size. Here, we produce an F₁ mapping population of 85 individuals from a cross between CP 1001 (dwarf commercial clone) and CP 96 (giant genotype), and use it to generate two linkage genetic maps comprising of 205 genetic markers (194 AFLP and 11 SSR markers). The female map (CP 1001) contains 122 markers over 19 linkage groups and the male map (CP 96) comprises 120 markers assembled over 23 linkage groups. The total map distance of the female map is 1050.7 cM representing around 68% genome coverage, whereas the male map spans 944.7 cM (64% coverage). The average map distance between markers is 8.6 cM in the female map and 7.9 cM in the male map. Homology between the two maps was established between 13 linkage groups of the female map and 14 of the male map using 46 bridging markers that include 11 SSR markers. These maps represent a platform from which to identify loci controlling economically important traits in this crop.     

Orientation and integration of the classical and molecular genetic maps of chromosome 11 in rice

Summary The classical genetic map and molecular map of rice chromosome 11 were oriented to facilitate the use of these maps for genetic studies and rice improvement. Three morphological markers (d-27, z-2, and la) were crossed to a rice breeding line, IRBB21, which has the Xa-21 gene for bacterial blight resistance. Three F₂ populations were analyzed with RFLP markers known to be located on chromosome 11. Segregation analysis of molecular markers and morphological markers was used to construct an RFLP map for each population. The recombination frequency between markers varied from population to population although the marker order on the maps was the same for all three populations. Based on a common set of markers mapped in the three populations, an integrated map was generated consisting of both RFLP and morphological markers. The genetic distance between markers on this map was determined by taking a weighted average of the data from the three populations. The oriented map serves as a bridge to understand the relationship between the classical and molecular linkage maps. Based on this information, the location of several genes on the classical map can be approximated with respect to RFLP markers without having to map them directly.     

An SSR-based molecular genetic map of cassava

Summary Microsatellites or simple sequence repeats (SSR) are the markers of choice for molecular genetic mapping and marker-assisted selection in many crop species. A microsatellite-based linkage map of cassava was drawn using SSR markers and a F₂ population consisting of 268 individuals. The F₂ population was derived from selfing the genotype K150, an early yielding genotype from an F₁ progeny from a cross between two non-inbred elite cassava varieties, TMS 30572 and CM 2177-2 from IITA and CIAT respectively. A set of 472 SSR markers, previously developed from cassava genomic and cDNA libraries, were screened for polymorphism in K150 and its parents TMS 30572 and CM 2177-2. One hundred and twenty two polymorphic SSR markers were identified and utilized for linkage analysis. The map has 100 markers spanning 1236.7 cM, distributed on 22 linkage groups with an average marker distance of 17.92 cM. Marker density across the genome was uniform. This is the first SSR based linkage map of cassava and represents an important step towards quantitative trait loci mapping and genetic analysis of complex traits in M. esculenta species in national research program and other institutes with minimal laboratory facilities. SSR markers reduce the time and cost of mapping quantitative trait loci (QTL) controlling traits of agronomic interest, and are of potential use for marker-assisted selection (MAS).     

Construction of high-density linkage map and identification of QTLs associated with resistance to black rot in radish (Raphanus sativus) by RAD sequencing

High-density marker-based QTL mapping can serve as an effective strategy to identify novel genomic information to facilitate crop improvement. In this study, we genotyped an F₂ population (KB12-1 × PP12-1) using a RAD-seq approach and constructed a high-density linkage map for radish. After a series of filtering procedures were performed, 17,124 SNPs and 3,336 indels with aa × bb genotyping were retained to obtain bin markers. Then, a linkage map comprising a total of 1,221 bin markers in nine linkage groups spanning 1,467.3 cM with an average marker interval of 1.2 cM was constructed. We evaluated the resistance of the F₂ mapping population to black rot using F₃ progeny, and two major QTLs related to black rot resistance were identified based on this map. Among these QTLs, qBRR2 on Chr.2 explained 26.97% of the phenotypic variation with a LOD score of 11.93, and qBRR7 on Chr.7 accounted for 27.06% of the phenotypic variation with a LOD score of 11.83. The additive effect of qBRR2 was positive (14.97); however, qBRR7 had the opposite effect (−11.99). The high-density linkage map and the major QTLs qBRR2 and qBRR7 provide new important information for disease resistance gene discovery and utilization in genetic improvement.     

AFLP-based STS markers closely linked to a fertility restoration locus (Rfm1) for cytoplasmic male sterility in barley

The Rfm1 gene restores the fertility of the msm1 and msm² male‐sterile cytoplasms in barley. Rfm1 is located on the short arm of chromosome 6H. To develop molecular markers tightly linked to Rfm1 for use in sophisticated marker‐assisted selection and map‐based cloning, an amplified fragment‐length polymorphism (AFLP) marker system with isogenic lines and a segregating BC₁F₁ population was used. Nine hundred primer combinations were screened and a linkage map was constructed around the Rfm1 locus by using 25 recombinant plants selected from 214 BC₁F₁ plants. Three AFLP markers were identified, e34m2, e46m19 and e48m17, linked to the locus. The most closely linked markers were e34m2, at 1.0 cM distally and e46m19, at 1.1 cM proximally. The two AFLP markers were converted to dominant STS markers. These markers should accelerate programmes for breeding restorer lines and will be useful for map‐based cloning.     

Mapping of the Vrn-B1 gene in Triticum aestivum using microsatellite markers

An F₂ population segregating for the dominant gene Vrn‐B1 was developed from the cross of the substitution line ‘Diamant/'Miro‐novskaya 808 5A’ and the winter wheat cultivar ‘Bezostaya 1′. Microsatellite markers (Xgwm and Xbarc) with known map locations on chromosome 5B of common wheat were used for mapping the gene Vrn‐B1. Polymorphism between parental varieties was observed for 28 out of 34 microsatellite markers (82%). Applying the quantitative trait loci mapping approach, the target gene was mapped on the long arm of chromosome 5B, closely linked to Xgwm408. The map position of Vrn‐B1 suggests that the gene is homoeologous to other vernalization response genes located on the homoeologous group 5 chromosomes of wheat, rye and barley.     

RFLP-based mapping of the Sec-2 and Sec-5 loci encoding 75K γ-secalins of rye

For mapping the Sec2 and Sec5 loci of rye which determine expression of 75K γ-secalins, a partial genetic map of chromosome 2R spanning 64 cM was constructed. The map was developed using an F₂ population of 103 plants from a cross between two inbred lines. Both loci were mapped distally on the short arm of chromosome 2R and clearly tagged in relation to 12 restriction fragment length polymorphism (RFLP) markers. The Sec2 locus was localized between the Xiag57 and Xpsr109a loci in an 11 cM interval. The Sec5 locus co-segregated to Xiag57 and was tightly linked to the Sec2 locus at a map distance of 0.5cM.     

Comparison and integration of genetic maps generated from F2 and BC1-type mapping populations in perennial ryegrass

Linkage maps of perennial ryegrass were constructed from F₂ and BC₁‐type populations using, predominantly, restriction fragment length polymorphism data based on heterologous probes used in mapping other grass species. The maps identified seven linkage groups, which covered a total of 515 cM (F₂) and 565 cM (BC₁). They were aligned using 38 loci identified in both populations (common loci) and a possible marker order for all mapped loci in either population was identified in an integrated map. The estimated recombination frequencies and map distances between adjacent common loci were compared between the two data sets and regions of heterogeneity identified. Overall, the common markers identified a map distance of 446 cM in the F₂ population and 327 cM in the BC₁ population, reflecting a higher recombination frequency in the former, although the difference was not evenly spread over the seven linkage groups.     

T1 locus in cotton is the candidate gene affecting lint percentage, fiber quality and spiny bollworm (Earias spp.) resistance

A genetic linkage map of chromosome 6 was constructed by using 270 recombinant inbred lines originated from an upland cotton cross (Yumian 1 × T586) F₂ population. The genetic map included one morphological (T₁) and 18 SSR loci, covering 96.2 cM with an average distance of 5.34 cM between two markers. Based on composite interval mapping (CIM), QTL(s) affecting lint percentage, fiber length, fiber length uniformity, fiber strength and spiny bollworm resistance (Earias spp.) were identified in the t₁ locus region on chromosome 6. The allele(s) originating from T586 of QTLs controlling lint percentage increased the trait phenotypic value while the alleles originating from Yumian 1 of QTLs affecting fiber length, fiber length uniformity, fiber strength and spiny bollworm resistance increased the trait phenotypic value.     

Development of EST-SSR markers and construction of a linkage map in faba bean (Vicia faba)

To develop a high density linkage map in faba bean, a total of 1,363 FBES (Faba bean expressed sequence tag [EST]-derived simple sequence repeat [SSR]) markers were designed based on 5,090 non-redundant ESTs developed in this study. A total of 109 plants of a ‘Nubaria 2’ × ‘Misr 3’ F₂ mapping population were used for map construction. Because the parents were not pure homozygous lines, the 109 F₂ plants were divided into three subpopulations according to the original F₁ plants. Linkage groups (LGs) generated in each subpopulation were integrated by commonly mapped markers. The integrated ‘Nubaria 2’ × ‘Misr 3’ map consisted of six LGs, representing a total length of 684.7 cM, with 552 loci. Of the mapped loci, 47% were generated from multi-loci diagnostic (MLD) markers. Alignment of homologous sequence pairs along each linkage group revealed obvious syntenic relationships between LGs in faba bean and the genomes of two model legumes, Lotus japonicus and Medicago truncatula. In a polymorphic analysis with ten Egyptian faba bean varieties, 78.9% (384/487) of the FBES markers showed polymorphisms. Along with the EST-SSR markers, the dense map developed in this study is expected to accelerate marker assisted breeding in faba bean.     

Detection of linkage disequilibrium QTLs controlling low-temperature growth and metabolite accumulations in an admixed breeding population of <Emphasis Type="BoldItalic">Leymus</Emphasis> wildryes

Low-temperature soluble carbohydrate accumulations are commonly associated with anthocyanin coloration, attenuated growth, and cold adaptation of cool-season grasses. A total of 647 AFLP markers were tested for associations with anthocyanin coloration, tiller formation, leaf formation, cumulative leaf length, percent soluble carbohydrate, and dry matter regrowth among replicated clones of an admixed Leymus wildrye breeding population evaluated in low-temperature growth chambers. The admixed breeding population was derived from a heterogeneous population of L. cinereus × L. triticoides F₁ hybrids, with two additional generations of open pollination. Two AFLP linkage maps, constructed from two full-sib mapping populations derived from the same F₁ hybrid population, were integrated to produce a framework consensus map used to examine the distribution of marker-trait associations in the admixed F₁OP₂ population. Thirty-seven linkage blocks, spanning 258 cM (13.6%) of the 1895 cM consensus map, contained 119 (50%) of the 237 markers showing at least one possible trait association (P < 0.05). Moreover, 28 (68%) of the 41 most significant marker-trait associations (P < 0.005) were located in 15 QTL linkage blocks spanning 112.9 cM (6%) of the linkage map. The coincidence of these 28 significant marker-trait associations, and many less significant associations, in 15 relatively small linkage blocks (0.6 cM to 21.3 cM) provides evidence of admixture linkage disequilibrium QTLs (ALD QTLs) in this heterogeneous breeding population. At least four of the remaining 13 putative marker-trait associations (P < 0.005) were located in genetic map regions lacking other informative markers. The complexity of marker-trait associations results from heterogeneity within and substantial divergence among the parental accessions.     

Development of an interspecific Vigna linkage map between Vigna umbellata (Thunb.) Ohwi & Ohashi and V. nakashimae (Ohwi) Ohwi & Ohashi and its use in analysis of bruchid resistance and comparative genomics

To facilitate transfer of bruchid resistance to azuki bean (Vigna angularis) from its relatives an interspecific mapping population was made between rice bean, V. umbellata, and the related wild species V. nakashimae. The V. umbellata parent is completely resistant and V. nakashimae is completely susceptible to the bruchid beetle pests, azuki bean weevil (Callosobruchus chinensis) and cowpea weevil (C. maculatus). There is very low cross compatibility between V. umbellata and azuki bean. Therefore, V. nakashimae, that crosses with both V. umbellata and V. angularis without the need for embryo rescue, is used as a bridging species. A genetic linkage map was constructed based on an interspecific F₂ mapping population between V. umbellata and V. nakashimae consisting of 74 plants. A total of 175 DNA marker loci (74 RFLPs and 101 SSRs) were mapped on to 11 linkage groups spanning a total length of 652 cM. Segregation distortion was observed but only three markers were not linked to any linkage group due to severe segregation distortion. This interspecific genome map was compared with the genome map of azuki bean. Of 121 common markers on the two maps, 114 (94.2%) were located on the same linkage groups in both maps. The marker order was highly conserved between the two genome maps. Fifty F₂ plants that produced sufficient seeds were used for quantitative trait locus (QTL) analysis and locating gene(s) for C. chinensis and C. maculatus resistance in V. umbellata. The resistance reaction of these F₂ plants differed between C. chinensis and C. maculatus. Both resistances were quantitatively inherited with no F₂ plants completely susceptible to C. chinensis or C. maculatus. One putative QTL for resistance to each of these bruchid species was located on different linkage groups. Other putative QTLs associated with resistance to both C. chinensis and C. maculatus were localized on the same linkage group 1. Linked markers associated with the bruchid‐resistant QTL will facilitate their transfer to azuki bean breeding lines.     

Construction of a genetic map of melon with molecular markers and horticultural traits, and localization of genes associated with ZYMV resistance

Abstract: A partial linkage map of melon was constructed from a cross between PI414723 and Dulce. Twenty-two SSR, 46RAPD, 2 ISSR markers and four horticultural markers [female flower form (a), Fusarium resistance, striped epicarp (st), and fruit flesh pH (pH)] were analyzed in an F₂/F₃ population to produce a map spanning 14 linkage groups. We report for the first time map positions for the st, a, and pH genes. One SSR marker was tightly linked to pH. Mapping the a gene for the female flower form to molecular linkage group 4 enabled the merging of the map of horticultural traits with the of molecular markers in this region. Using the 22 SSR markers of this map, two of the three postulated ZYMV resistance genes were located using a BC₁ population (PI414723 recurrent parent). One SSR marker was tightly linked to a ZYMV resistance gene, designated Zym-1. This revised version was published online in August 2006 with corrections to the Cover Date.     

Development of tomato SSR markers from anchored BAC clones of chromosome 12 and their application for genetic diversity analysis and linkage mapping

In this study, we developed a total of 37 simple sequence repeat (SSR) markers from 11 bacterial artificial chromosome (BAC) clone sequences anchored on chromosome 12 of tomato available at Solanaceae Genomics Network. These SSR markers could group a set of 16 tomato genotypes comprising of Solanum lycopersicum, S. pimpinellifolium, S. habrochaites, and S. pennellii unambiguously according to their known species status. Clear subgroups of genotypes within S. lycopersicum were also observed. A subset of 16 SSR markers representing the 11 BAC clones was used for developing genetic linkage maps of three interspecific F₂ populations produced from the crosses involving a common S. lycopersicum parent (CLN2498E) with S. pennellii (LA1940), S. habrochaites (LA407) and S. pimpinellifolium (LA1579). The length of the genetic linkage maps were 112.5 cM, 109.3 cM and 114.1 cM, respectively. Finally, an integrated genetic linkage map spanning a total length of 118.7 cM was developed. The reported SSR markers are uniformly distributed on chromosome 12 and would be useful for genetic diversity and mapping studies in tomato.     

Molecular mapping of genes involved in root hair formation in barley

In the presented study, the existing AFLP and SSR maps of barley were used to find chromosomal position of four genes controlling different stages of root hair development. Four barley mutants were used in the analysis: the root hairless mutant rhl1.b, mutant rhp1.b with root hair development blocked at the initial bulge formation, mutant rhi1.a with irregular pattern of sparsely located root hairs and mutant rhs1.a with very short root hairs. Each mutant was crossed with parents of ‘Steptoe’/‘Morex’ mapping population and F₂ progenies of crosses: mutant × ‘Steptoe’ and mutant × ‘Morex’ were analyzed for segregation of root hair phenotype and polymorphic AFLP and SSR markers. It was possible to map all the analyzed genes on barley chromosomes: rhl1 gene on the short arm of chromosome 7H, rhp1 gene on chromosome 1H, rhs1 locus in the pericentromeric region of chromosome 5H and rhi1 gene on the long arm of chromosome 6H. Subsequently, the Bulk Segregant Analysis and AFLP technique were used for saturation of the identified regions with new markers. The joint maps were constructed using as common points the SSR markers located in the target regions. Linkage maps of the regions around the four genes involved in the root hair formation in barley were composed of 8–11 markers and spanned over 16.1–49.0 cM. The distances between localized genes and the closest markers ranged from 1.0 to 3.8 cM. The identified chromosomal locations of genes can be used for their fine mapping and future map-based cloning.