Genetic localization of a bruchid resistance gene and its relationship to insecticidal cyclopeptide alkaloids, the vignatic acids, in mungbean ( Vigna radiata L. Wilczek)

Bruchid resistance, controlled by a single dominant gene (Br) in a wild mungbean accession (TC1966), has been incorporated into cultivated mungbean (Vigna radiata). The resistance gene simultaneously confers inhibitory activity against the bean bug, Riptortus clavatus Thunberg (Hemiptera: Alydidae). The resultant isogenic line (BC₂₀ generation) was characterized by the presence of a group of novel cyclopeptide alkaloids, called vignatic acids. A linkage map was constructed for Br and the vignatic acid gene (Va) using restriction fragment length polymorphism (RFLP) markers and a segregating BC₂₀F₂ population. By screening resistant and susceptible parental lines with 479 primers, eight randomly amplified polymorphic DNA (RAPD) markers linked to Br were identified and cloned for use as RFLP probes. All eight RAPD-based markers, one mungbean, and four common bean genomic clones were effectively integrated around Br within a 3.7-cM interval. Br was mapped to a 0.7-cM segment between a cluster consisting of six markers and a common bean RFLP marker, Bng110. The six markers are closest to the bruchid resistance gene, approximately 0.2 cM away. The vignatic acid gene, Va, cosegregated with bruchid resistance. However, one individual was identified in the BC₂₀F₂ population that retained vignatic acids in spite of its bruchid susceptibility. Consequently, Va was mapped to a single locus at the same position as the cluster of markers and 0.2 cM away from Br. These results suggest that the vignatic acids are not the principal factors responsible for bruchid resistance in V. radiata but will facilitate the use of map-based cloning strategies to isolate the Br gene. Received: 20 November 1997 / Accepted: 6 January 1998     

Genetic localization of a bruchid resistance gene and its relationship to insecticidal cyclopeptide alkaloids, the vignatic acids, in mungbean (Vigna radiata L. Wilczek)

Bruchid resistance, controlled by a single dominant gene (Br) in a wild mungbean accession (TC1966), has been incorporated into cultivated mungbean (Vigna radiata). The resistance gene simultaneously confers inhibitory activity against the bean bug, Riptortus clavatus Thunberg (Hemiptera: Alydidae). The resultant isogenic line (BC₂₀ generation) was characterized by the presence of a group of novel cyclopeptide alkaloids, called vignatic acids. A linkage map was constructed for Br and the vignatic acid gene (Va) using restriction fragment length polymorphism (RFLP) markers and a segregating BC₂₀F₂ population. By screening resistant and susceptible parental lines with 479 primers, eight randomly amplified polymorphic DNA (RAPD) markers linked to Br were identified and cloned for use as RFLP probes. All eight RAPD-based markers, one mungbean, and four common bean genomic clones were effectively integrated around Br within a 3.7-cM interval. Br was mapped to a 0.7-cM segment between a cluster consisting of six markers and a common bean RFLP marker, Bng110. The six markers are closest to the bruchid resistance gene, approximately 0.2?cM away. The vignatic acid gene, Va, cosegregated with bruchid resistance. However, one individual was identified in the BC₂₀F₂ population that retained vignatic acids in spite of its bruchid susceptibility. Consequently, Va was mapped to a single locus at the same position as the cluster of markers and 0.2?cM away from Br. These results suggest that the vignatic acids are not the principal factors responsible for bruchid resistance in V. radiata but will facilitate the use of map-based cloning strategies to isolate the Br gene.     

A genetic linkage map of azuki bean constructed with molecular and morphological markers using an interspecific population (Vigna angularis x V. nakashimae)

A genetic linkage map of azuki bean (Vigna angularis) was constructed with molecular and morphological markers using an F₂ population of an interspecific cross between azuki bean and its wild relative, V. nakashimae. In total, 132 markers (108 RAPD, 19 RFLP and five morphological markers) were mapped in 14 linkage groups covering 1250 cM; ten remained unlinked. The clusters of markers showing distorted segregation were found in linkage groups 2, 8 and 12. By comparing the azuki linkage map with those of mungbean and cowpea, using 20 RFLP common markers, some sets of the markers were found to belong to the same linkage groups of the respective maps, indicating that these linkage blocks are conserved among the three Vigna species. This map provides a tool for markerassisted selection and for studies of genome organization in Vigna species.     

Characterization and mapping of <Emphasis Type="Italic">Rpi1</Emphasis>, a gene that confers dominant resistance to stalk rot in maize

The maize inbred lines 1145 (resistant) and Y331 (susceptible), and the F₁, F₂ and BC₁F₁ populations derived from them were inoculated with the pathogen Pythium inflatum Matthews, which causes stalk rot in Zea mays. Field data revealed that the ratio of resistant to susceptible plants was 3:1 in the F₂ population, and 1:1 in the BC₁F₁population, indicating that the resistance to P. inflatum Matthews was controlled by a single dominant gene in the 1145×Y331 cross. The gene that confers resistance to P. inflatum Matthews was designated Rpi1 for resistance to P. inflatum) according to the standard nomenclature for plant disease resistance genes. Fifty SSR markers from 10 chromosomes were first screened in the F₂ population to find markers linked to the Rpi1 gene. The results indicated that umc1702 and mmc0371 were both linked to Rpi1, placing the resistance gene on chromosome 4. RAPD (randomly amplified polymorphic DNA) markers were then tested in the F₂population using bulked segregant analysis (BSA). Four RAPD products were found to show linkage to the Rpi1 gene. Then 27 SSR markers and 8 RFLP markers in the region encompassing Rpi1 were used for fine-scale mapping of the resistance gene. Two SSR markers and four RFLP markers were linked to the Rpi1 gene. Finally, the Rpi1 gene was mapped between the SSR markers bnlg1937 and agrr286 on chromosome 4, 1.6 cM away from the former and 4.1 cM distant from the latter. This is the first time that a dominant gene for resistance to maize stalk rot caused by P. inflatum Matthews has been mapped with molecular marker techniques.     

Identification of an RFLP marker tightly linked to theHt1 gene in maize

Summary We have identified tight linkage of an RFLP marker to theHt1 gene of maize that confers resistance to the fungal pathogenHelminthosporium turcicum race 1. This was accomplished by the use of four pairs of near isogenic lines (NILs; B73, A619, W153R, and CM105), each differing by the presence or the absence of the geneHt1. SinceHt1 maps to chromosome 2, 26 clones already mapped to this chromosome were labeled and probed against Southern blots of these NILs DNA digested with three restriction enzymes:EcoRI,BamHI, andHindIII. Six markers exhibited an RFLP for at least one pair of NILs. Presumptive linkage was further tested by analyzing the segregation of five of the six markers (one was monomorphic in the cross studied) and resistance toH. turcicum race 1 on 95 F₂ individuals from the cross DF20 × LH146Ht. The results indicate a tight linkage between one of the DNA markers,UMC150B, and theHt1 gene.     

绿豆抗豆象遗传的初步研究

绿豆象(Callos0bruchun chinensis L.)是豇豆属豆类作物重要的仓库害虫.本研究通过抗豆象杂交育种后代VC1973A/TC1966 F1、F2和VC1973A/(VC1973A/TC1966 F2)BC1F1及TC1966/(VC1973A/TC1966 F2)BC1F1分离群体的遗传分析,发现绿豆抗豆象性状符合31的遗传分离规律,证明绿豆对豆象的抗性由1对显性单基因(Aa)控制,抗虫性为显性(A),感虫为隐性(a).     

Comparative genome analysis of mungbean (Vigna radiata L. Wilczek) and cowpea (V. unguiculata L. Walpers) using RFLP mapping data

Genome relationships between mungbean (Vigna tradiata) and cowpea (V. Unguiculata) based on the linkage arrangement of random genomic restriction fragment length polymorphism (RFLP) markers have been investigated. A common set of probes derived from cowpea, common bean (Phaseolus vulgaris), mungbean, and soybean (Glycine max) PstI genomic libraries were used to construct the genetic linkage maps. In both species, a single F₂ population from a cross between an improved cultivar and a putative wild progenitor species was used to follow the segregation of the RFLP markers. Approximately 90% of the probes hybridized to both mungbean and cowpea DNA, indicating a high degree of similarity in the nucleotide sequences among these species. A higher level of polymorphism was detected in the mungbean population (75.7%) than in the cowpea population (41.2%). Loci exhibiting duplications, null phenotypes, and distorted segregation ratios were detected in both populations. Random genomic DNA RFLP loci account for about 89% of the currently mapped markers with a few cDNA and RAPD markers added. The current mungbean map is comprised of 171 loci/loci clusters distributed in 14 linkage groups spanning a total of 1570cM. On the other hand, 97 markers covered 684 cM and defined 10 linkage groups in the current cowpea map. The mungbean and cowpea genomes were compared on the basis of the copy number and linkage arrangement of 53 markers mapped in common between the two species. Results indicate that nucleotide sequences are conserved, but variation in copy number were detected and several rearrangements in linkage orders appeared to have occurred since the divergence of the two species. Entire linkage groups were not conserved, but several large linkage blocks were maintained in both genomes.     

Identification and mapping on chromosome 9 of RAPD markers linked to Sw-5 in tomato by bulked segregant analysis

Bulked segregant analysis was used to identify random amplified polymorphic DNA (RAPD) markers linked to the Sw-5 gene for resistance to tomato spotted wilt virus (TSWV) in tomato. Using two pools of phenotyped individuals from one segregating population, we identified four RAPD markers linked to the gene of interest. Two of these appeared tightly linked to Sw-5, whereas another, linked in repulsion phase, enabled the identification of heterozygous and susceptible plants. After linkage analysis of an F₂ population, the RAPD markers were shown to be linked to Sw-5 within a distance of 10.5 cM. One of the RAPD markers close to Sw-5 was used to develop a SCAR (sequence characterized amplified region) marker. Another RAPD marker was stabilized into a pseudo-SCAR marker by enhancing the specificity of its primer sequence without cloning and sequencing. RAPD markers were mapped to chromosome 9 on the RFLP tomato map developed by Tanksley et al. (1992). The analysis of 13 F₃ families and eight BC₂ populations segregating for resistance to TSWV confirmed the linkage of the RAPD markers found. These markers are presently being used in marker-assisted plant breeding.     

Identification and mapping of microsatellite markers linked to a root-knot nematode resistance gene (rkn1) in Acala NemX cotton (Gossypium hirsutum L.)

Host-plant resistance is the most economic and effective strategy for root-knot nematode (RKN) Meloidogyne incognita control in cotton (Gossypium hirsutum L.). Molecular markers linked to resistance are important for incorporating resistance genes into elite cultivars. To screen for microsatellite markers (SSR) closely linked to RKN resistance in G. hirsutum cv. Acala NemX, F₁, F₂, BC₁F₁, and F_2:7 recombinant inbred lines (RILs) from intraspecific crosses and an F₂ from an interspecific cross with G. barbadense cv. Pima S-7 were used. Screening of 284 SSR markers, which cover all the known identified chromosomes and most linkage groups of cotton, was performed by bulked segregant analysis, revealing informative SSRs. The informative SSRs were then mapped on the above populations. One co-dominant SSR marker CIR316 was identified tightly linked to a major resistance gene (designated as rkn1), producing amplified DNA fragments of approximately 221 bp (CIR316a) and 210 bp (CIR316c) in Acala NemX and susceptible Acala SJ-2, respectively. The linkage between CIR316a marker and resistance gene rkn1 in Acala NemX had an estimated distance of 2.1–3.3 cM depending on the population used. Additional markers, including BNL1231 with loose linkage to rkn1 (map distance 25.1–27.4 cM), BNL1066, and CIR003 allowed the rkn1 gene to be mapped to cotton linkage group A03. This is the first report in cotton with a closely linked major gene locus determining nematode resistance, and informative SSRs may be used for marker-assisted selection.     

RFLP and RAPD mapping of the sunflower Pl1 locus for resistance to Plasmopara halstedii race 1

The Pl1 locus in sunflower, Helianthus annuus L., conferring resistance to downy mildew, Plasmopara halstedii, race 1 has been located in linkage group 1 of the consensus RFLP map of the cultivated sunflower. Bulked segregant analyses were used on 135 plants of an F₂ progeny from a cross between a downy mildew susceptible line, GH, and RHA266, a line carrying Pl1. Two RFLP markers and one RAPD marker linked to the Pl1 locus have been identified. The RFLP markers are located at 5.6 cM and 7.1 cM on either side of Pl1. The RAPD marker is situated at 43.7 cM from Pl1. The significance and applications of these markers in sunflower breeding are discussed.     

A barley RFLP map: alignment of three barley maps and comparisons to Gramineae species

Several gene linkage maps have been produced for cultivated barley. We have produced a new linkage map for barley, based on a cross between Hordeum vulgare subsp. spontaneum and Hordeum vulgare subsp. vulgare (Hvs x Hvv), having a higher level of polymorphism than most of the previous barley crosses used for RFLP mapping. Of 133 markers mapped in the Hvs x Hvv F₂ population, 69 were previously mapped on other barley maps, and 26 were mapped in rice, maize, or wheat. Two known gene clones were mapped as well as two morphological markers. The distributions of previously mapped markers were compared with their respective barley maps to align the different maps into one consensus map. The distributions of common markers among barley, wheat, rice and maize were also compared, indicating colinear linkage groups among these species.To be considered dual first authorsPublished with the approval of the Director of the Colorado State University/Agricultural Experiment Station.     

Genetic analysis and gene mapping of a rice few-tillering mutant in early backcross populations ( Oryza sativa L.)

A rice mutant,G069, characteristic of few tiller numbers, was found in anther culture progeny from theF ₁ hybrid between anindica-japonica cross, Gui630×02428. The mutant has another two major features: delayed tillering development and yellowing apex and margin on the mature leaves. As a donor parent,G069 was further backcrossed with the recurrent parent,02428, for two turns to develop aBC ₂F₂ population. Genetic analysis in theBC ₂F₂ population showed that the traits of few-tillering and yellowing apex and margin on the mature leaves were controlled by one recessive gene. A pool of equally mixed genomic DNA, from few-tillering individual plants inBC ₂F₂, was constructed to screen polymorphism with simple sequence repeat (SSR) markers in comparison with the02428 genome. One SSR marker and three restriction fragment length polymorphism (RFLP) markers were found possibly linked with the recessive gene. By using these markers, the gene of few-tillering was mapped on chromosome 2 between RFLP marker C424 and S13984 with a genetic distance of 2.4 cM and 0.6 cM, respectively. The gene is designatedft1.     

Identification and mapping of AFLP markers linked to peanut (<Emphasis Type="Italic">Arachis hypogaea</Emphasis> L.) resistance to the aphid vector of groundnut rosette disease

Groundnut rosette disease is the most destructive viral disease of peanut in Africa and can cause serious yield losses under favourable conditions. The development of disease-resistant cultivars is the most effective control strategy. Resistance to the aphid vector, Aphis craccivora, was identified in the breeding line ICG 12991 and is controlled by a single recessive gene. Bulked segregant analysis (BSA) and amplified fragment length polymorphism (AFLP) analysis were employed to identify DNA markers linked to aphid resistance and for the development of a partial genetic linkage map. A F_2:3 population was developed from a cross using the aphid-resistant parent ICG 12991. Genotyping was carried out in the F₂ generation and phenotyping in the F₃ generation. Results were used to assign individual F₂ lines as homozygous-resistant, homozygous-susceptible or segregating. A total of 308 AFLP (20 EcoRI+3/MseI+3, 144 MluI+3/MseI+3 and 144 PstI+3/MseI+3) primer combinations were used to identify markers associated with aphid resistance in the F_2:3 population. Twenty putative markers were identified, of which 12 mapped to five linkage groups covering a map distance of 139.4 cM. A single recessive gene was mapped on linkage group 1, 3.9 cM from a marker originating from the susceptible parent, that explained 76.1% of the phenotypic variation for aphid resistance. This study represents the first report on the identification of molecular markers closely linked to aphid resistance to groundnut rosette disease and the construction of the first partial genetic linkage map for cultivated peanut.     

Identification and mapping of pm2026: a recessive powdery mildew resistance gene in an einkorn (Triticum monococcum L.) accession

Triticum monococcum accession TA2026 showed resistance to wheat powdery mildew. To identify the resistance gene and transfer it to common wheat, genetic analysis and molecular mapping were conducted using an F₂ population and derived F₃ families from the cross of TA2026 × M389. The results indicated that TA2026 possessed a recessive powdery mildew resistance gene. This gene was mapped to the terminal portion of chromosome 5A^mL and flanked by SSR marker loci Xcfd39 and Xgwm126. Eight RFLP markers previously mapped to the terminal chromosome 5A^mL were converted into STS markers. Three loci, detected by MAG1491, MAG1493 and MAG1494, the STS markers derived from RFLP probes CDO1312, PSR164 and PSR1201, respectively, were linked to this resistance gene with Xmag1493 only 0.9 cM apart from it. In addition, the STS marker MAG2170 developed from the tentative consensus wheat cDNA encoding the Mlo-like protein identified a locus co-segregating with Xmag1493. This is the first recessive powdery mildew resistance gene identified on chromosome 5A^m, and is temporarily designated pm2026. We have successfully transferred it to a tetraploid background, and this resistance stock will now be used as the bridge parent for its transfer to common wheat.     

Identification and mapping of resistance gene analogs and a white rust resistance locus in<Emphasis Type="Italic"> Brassica rapa</Emphasis> ssp.<Emphasis Type="Italic"> oleifera</Emphasis>

The objective of this investigation was to tag a locus for white rust resistance in a Brassica rapa ssp. oleifera F₂ population segregating for this trait, using bulked segregant analysis with random amplified polymorphic DNA (RAPD) markers, linkage mapping and a candidate gene approach based on resistance gene analogs (RGAs). The resistance source was the Finnish line Bor4109. The reaction against white rust races 7a and 7v was scored in 20 seedlings from each self-pollinated F₂ individual. The proportion of resistant plants among these F₃ families varied from 0 to 67%. Bulked segregant analysis did not reveal any markers linked with resistance and, therefore, a linkage map with 81 markers was created. A locus that accounted for 18.4% of the variation in resistance to white rust was mapped to linkage group (LG) 2 near the RAPD marker Z19a. During the study, a bacterial resistance gene homologous to Arabidopsis RPS2 and six different RGAs were sequenced. RPS2 and five of the RGAs were mapped to linkage groups LG1, LG4 and LG9. Unfortunately, none of the RGAs could be shown to be associated with white rust resistance.Communicated by H.C. BeckerThe nucleotide sequence data reported has been deposited in the Genbank under the accession numbers AF315081–AF315087.     

Molecular markers linked with bruchid resistance in Vigna radiata var. Sublobata and their validation

Bruchid, Callosobruchus chinensis (L.) is an important pest of Vigna radiata during storage. RFLP and PCR based markers identified, linked with bruchid resistance gene in wild accession of greengram (V. radiata var. Sublobata) either collected from Madagaskar or Australia. Whether these markers will be useful for marker assisted introgression of bruchid resistance gene from the Indian accession into the existing cultivars are not known. Here, we employed two STS based markers which were found earlier, to be linked with bruchid resistance gene in Australian accession ACC41. Only one primer pair, STSbr1 showed polymorphism among Indian Sublobata accession (Sub2) and other twelve green gram cultivars. Analysis of 113 segregating lines (F₆) of a cross between a popular cultivar of West Bengal, B1 and Sub2 showed a cent percent co-segregation of resistant locus with the polymorphic fragment. STSbr1 behave as a dominant marker among Indian genotypes although it has been shown earlier a co-dominant banding pattern between ACC41 and other Australian Susceptible cultivars. Other STS marker, STSbr2, does not produce any polymorphic fragment among Sub2 and 18 greengram genotypes. STSbr1 employed in screening of 50 green gram accessions and found high efficiency in screening of bruchid resistant genotypes also. So STSbr1 will be useful for marker assisted selection and germplasm screening for development of bruchid resistant greengram.     

Molecular identification of powdery mildew resistance genes in common wheat (Triticum aestivum L.)

RFLP markers for the wheat powdery mildew resistance genes Pm1 and Pm2 were tagged by means of near-isogenic lines. The probe Whs178 is located 3 cM from the Pm1 gene. For the powdery mildew resistance gene Pm2, two markers were identified. The linkage between the Pm2 resistance locus and one of these two probes was estimated to be 3 cM with a F₂ population. Both markers can be used to detect the presence of the corresponding resistance gene in commercial cultivars. Bulked segregant analysis was applied to identify linkage disequillibrium between the resistance gene Pm18 and the abovementioned marker, which was linked to this locus at a distance of 4 cM. Furthermore, the RAPD marker OPH-11₁₉₀₀ (5-CTTCCGCAGT-3) was selected with pools created from a population segregating for the resistance of Trigo BR 34. The RAPD marker was mapped about 13 cM from this resistance locus.     

The Ry-fsto gene from Solanum stoloniferum for extreme resistant to Potato virus Y maps to potato chromosome XII and is diagnosed by PCR marker GP122718 in PVY resistant potato cultivars

A novel locus for extreme resistance to Potato virus Y (PVY), Ry-f_sto, was identified on potato chromosome XII. The gene Ry-f_sto has been introgressed from the wild potato species Solanum stoloniferum. Inheritance of Ry-f_sto in the tetraploid potato population Rysto was consistent with the model of a single, dominant gene. Bulked segregant analysis identified an ISSR (inter-simple sequence repeat) marker UBC 857₉₈₀ linked to Ry-f_sto. This marker mapped to linkage group XII of a reference potato RFLP (restriction fragment length polymorphism) map. Chromosome XII specific RFLP markers were converted into PCR-based STS and CAPS markers and tested for linkage with Ry-f_sto in the population Rysto. CAPS marker GP122₇₁₈ was tightly linked to the resistance gene and was successfully used to identify Polish and German cultivars expressing extreme resistance to PVY. This indicates that the source of Ry-f_sto has been widely utilized in various potato breeding programs and can be monitored by a diagnostic marker in marker-assisted selection.     

Mapping of loci involved in quantitatively inherited resistance to the potato cyst-nematode Globodera rostochiensis pathotype Ro1

We report the identification and mapping of two quantitative trait loci (QTLs) of Solanum spegazzinii BGRC, accession 8218-15, involved in resistance to the potato cyst-nematode Globodera rostochiensis pathotype Ro1, by means of restriction fragment length polymorphisms (RFLPs). For this purpose we crossed a susceptible diploid S. tuberosum with the resistant S. spegazzinii, and tested the F₁ population for resistance to the Ro1 pathotype. Since the F₁ segregated for the resistance, the S. spegazzinii parent was concluded to be heterozygous at the nematode resistance loci. For the mapping of the resistance loci we made use of RFLP markers segregating for S. spegazzinii alleles in the F₁. One hundred and seven RFLP markers were tested in combination with four different restriction enzymes; 29 of these displayed a heterozygous RFLP pattern within S. spegazzinii and were used for mapping. Analysis of variance (ANOVA) was applied to test the association of the RFLP patterns of these markers with nematode resistance. Two QTLs involved in disease resistance to Globodera rostochiensis pathotype Ro1 were identified and mapped to chromosomes 10 and 11 respectively.     

Linkage analysis of er-1, a recessive Pisum sativum gene for resistance to powdery mildew fungus (Erysiphe pisi D.C.)

Linkage analysis was used to determine the genetic map location of er-1, a recessive gene conditioning resistance to powdery mildew, on the Pisum sativum genome. Genetic linkage was demonstrated between er-1 and linkage group 6 markers after analyzing the progeny of two crosses, an F₂ population and a set of recombinant inbred lines. The classes of genetic markers surrounding er-1 include RFLP, RAPD and allozyme markers as well as the morphological marker Gty. A RAPD marker tightly linked to er-1 was identified by bulked segregant analysis. After DNA sequence characterization, specific PCR primers were designed to convert this RAPD marker into a sequence characterized amplified region (SCAR).