Marker enrichment and high-resolution map of the segment of potato chromosome VII harbouring the nematode resistance gene Gro1

The dominant allele Gro1 confers on potato resistance to the root cyst nematode Globodera rostochiensis. The Gro1 locus has been mapped to chromosome VII on the genetic map of potato, using RFLP markers. This makes possible the cloning of Gro1 based on its map position. As part of this strategy we have constructed a high-resolution genetic map of the chromosome segment surrounding Gro1, based on RFLP, RAPD and AFLP markers. RAPD and RFLP markers closely linked to Gro1 were selected by bulked segregant analysis and mapped relative to the Gro1 locus in a segregating population of 1105 plants. Three RFLP and one RAPD marker were found to be inseparable from the Gro1 locus. Two AFLP markers were identified that flanked Gro1 at genetic distances of 0.6 cM and 0.8 cM, respectively. A genetic distance of 1 cM in the Gro1 region corresponds to a physical distance of ca. 100 kb as estimated by long-range restriction analysis. Marker-assisted selection for nematode resistance was accomplished in the course of constructing the high-resolution map. Plants carrying the resistance allele Gro1 could be distinguished from susceptible plants by marker assays based on the polymerase chain reaction (PCR).     

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.     

Identification of RFLP markers closely linked to the H1 gene conferring resistance to Globodera rostochiensis in potato

Summary Resistance to the root cyst nematode Globodera rostochiensis is an agronomic trait that is at present incorporated into most new potato varieties. Major dominant genes are available that originate from wild and cultivated Solanum species closely related to the cultivated European potato (Solanum tuberosum ssp. tuberosum). One of those genes, H1, from S. Tuberosum ssp. andigena, was mapped to a distal position on potato chromosome V using restriction fragment length polymorphism (RFLP) markers. The H1 locus segregates independently from Gro1, a second dominant gene presumably from S. Spegazzinii that confers resistance to G. Rostochiensis and which has been mapped to chromosome VII. One marker, CP113, was linked without recombination to the H1 locus.     

Localization by restriction fragment length polymorphism mapping in potato of a major dominant gene conferring resistance to the potato cyst nematode Globodera rostochiensis

Summary A major dominant locus conferring resistance against several pathotypes of the root cyst nematode Globodera rostochiensis was mapped on the linkage map of potato using restriction fragment length polymorphism (RFLP) markers. The assessment of resistance versus susceptibility of the plants in the experimental population considered was based on an in vivo (pot) and an in vitro (petri dish) test. By linkage to nine RFLP markers the resistance locus Gro1 was assigned to the potato linkage group IX which is homologous to the tomato linkage group 7. Deviations from the additivity of recombination frequencies between Gro1 and its neighbouring markers in the pot test led to the detection of a few phenotypic misclassifications of small plants with poor root systems that limited the observation of cysts on susceptible roots. Pooled data from both tests provided better estimates of recombination frequencies in the linkage interval defined by the markers flanking the resistance locus.     

Marker enrichment and high-resolution map of the segment of potato chromosome VII harbouring the nematode resistance gene Gro1

The dominant allele Gro1 confers on potato resistance to the root cyst nematode Globodera rostochiensis. The Gro1 locus has been mapped to chromosome VII on the genetic map of potato, using RFLP markers. This makes possible the cloning of Gro1 based on its map position. As part of this strategy we have constructed a high-resolution genetic map of the chromosome segment surrounding Gro1, based on RFLP, RAPD and AFLP markers. RAPD and RFLP markers closely linked to Gro1 were selected by bulked segregant analysis and mapped relative to the Gro1 locus in a segregating population of 1105 plants. Three RFLP and one RAPD marker were found to be inseparable from the Gro1 locus. Two AFLP markers were identified that flanked Gro1 at genetic distances of 0.6 cM and 0.8 cM, respectively. A genetic distance of 1 cM in the Gro1 region corresponds to a physical distance of ca. 100 kb as estimated by long-range restriction analysis. Marker-assisted selection for nematode resistance was accomplished in the course of constructing the high-resolution map. Plants carrying the resistance allele Gro1 could be distinguished from susceptible plants by marker assays based on the polymerase chain reaction (PCR).     

Quantitatively-inherited resistance toGlobodera pallida is dominated by one major locus inSolanum spegazzinii

A high level of resistance toGlobodera pallida pathotypes Pa2 and Pa3 exists inSolanum spegazzinii, a wild relative of potato (S. tuberosum ssp.tuberosum). Here we report the mapping of loci involved in quantitatively-inherited nematode resistance with the use of RFLPs. One major locus,Gpa, was mapped on chromosome 5 and two minor loci on chromosomes 4 and 7 ofS. spegazzinii. Additionally, the contribution of the susceptible parent to nematode resistance was determined. TheGpa locus was solely responsible for the high resistance level found in the segregating population. However, the RFLP marker closely linked to this resistance locus showed a distorted segregation, with a shortage of plants having the resistance linked allele. Our results indicate that a prediction of the genetic constitution of a quantitative trait based solely on phenotypic observations can lead to erroneous conclusions.     

A QTL for broad-spectrum resistance to cyst nematode species (Globodera spp.) maps to a resistance gene cluster in potato

 Broad-spectrum resistance in potato to the potato cyst nematode (PCN) species Globodera rostochiensis and G. pallida is commonly regarded as a polygenically inherited trait. Yet, by use of QTL analysis and a selected set of PCN populations, resistance to both PCN species could be ascribed to the action of locus Grp1. Grp1 confers major resistance to G. rostochiensis line Ro₅-22 and G. pallida population Pa₂-D383 and partial resistance to G. pallida population Pa₃-Rookmaker. Grp1 was mapped on chromosome 5 using previously characterized AFLP markers. Cleaved amplified polymorphic sequence (CAPS) markers available for RFLP loci GP21 and GP179 revealed that Grp1 maps on a genomic region harboring other resistance factors to viral, fungal and nematodal pathogens. The present data indicate that Grp1 is a compound locus which contains multiple genes involved in PCN resistance. Received: 10 September 1997 / Accepted: 6 October 1997     

Mapping of the cyst nematode resistance locus Gpa2 in potato using a strategy based on comigrating AFLP markers

 The nematode resistance locus Gpa2 was mapped on chromosome 12 of potato using information on the genomic positions of 733 known AFLP markers. The minimum number of AFLP primer combinations required to map Gpa2 was three. This demonstrates that a reference collection of potato AFLP markers may be a valuable tool for mapping studies in potato. By use of RFLP probes, Gpa2 was more precisely mapped at the distal end of chromosome 12. Gpa2 confers resistance to a distinct group of populations of the potato cyst nematode Globodera pallida and originates from the same potato accession as locus H1, conferring resistance to pathotype Ro₁ of G. rostochiensis. This study shows that these two nematode resistance loci are unlinked and that Gpa2 is linked to the Rx1 locus conferring resistance to potato virus X. The efficiency of AFLPs for genetic mapping of a highly heterozygous crop like potato is discussed and compared with the RFLP technique. Received: 24 February 1997/Accepted: 2 May 1997     

Mapping of QTLs involved in nematode resistance, tuber yield and root development in Solanum sp.

A backcross population, derived from the cross (S. tuberosumxS. spegazzinii)xS. tuberosum was used to map QTLs involved in nematode resistance, tuber yield and root development. Complete linkage maps were available for the interspecific hybrid parent as well as the S. tuberosum parent, and interval mapping for all traits was performed for both. Additionally, the intra- and inter-locus interactions of the QTLs were examined. The Gro1.2 locus, involved in resistance to G. rostochiensis pathotype Ro1, that was previously mapped in the S. tuberosumxS. spegazzinii F₁ population, was located more precisely on chromosome 10. A new resistance locus, Gro1.4, also conferring resistance to G. rostochiensis pathotype Ro1, was found on chromosome 3. Different alleles of this locus originating from both parents contributed to the resistant phenotype, indicating multiallelism at this locus. No interlocus interactions were observed between these two resistance loci. For resistance to G. pallida no QTLs were detected. One minor QTL involved in tuber yield was located on chromosome 4. Two QTLs involved in root development and having large effects were mapped on chromosomes 2 and 6 and an epistatic interaction was found between these two loci.     

QTL analysis of potato tuber dormancy

The potential loss of chemical sprout inhibitors because of public concern over the use of pesticides underscores the desirability of breeding for long dormancy of potato (Solanum tuberosum L.) tubers. Quantitative trait locus (QTL) analyses were performed in reciprocal backcrosses between S. tuberosum and S. berthaultii toward defining the complexity of dormancy. S. berthaultii is a wild Bolivian species characterized by a short-day requirement for tuberization, long tuber dormancy, and resistance to several insect pests. RFLP alleles segregating from the recurrent parents as well as from the interspecific hybrid were monitored in two segregating progenies. We detected QTLs on nine chromosomes that affected tuber dormancy, either alone or through epistatic interactions. Alleles from the wild parent promoted dormancy, with the largest effect at a QTL on chromosome 2. Long dormancy appeared to be recessive in the backcross to S. berthaultii (BCB). In BCB the additive effects of dormancy QTLs accounted for 48% of the measured phenotypic variance, and adding epistatic effects to the model explained only 4% more. In contrast, additive effects explained only 16% of the variance in the backcross to S. tuberosum (BCT), and an additional 24% was explained by the inclusion of epistatic effects. In BCB variation at all QTLs detected was associated with RFLP alleles segregating from the hybrid parent; in BCT all QTLs except for two found through epistasis were detected through RFLP alleles segregating from the recurrent parent. At least three dormancy QTLs mapped to markers previously found to be associated with tuberization in these crosses.Paper number 55 of the Department of Fruit and Vegetable Science, Cornell University     

Segregation analysis and RFLP mapping of the R1 and R3 alleles conferring race-specific resistance to Phytophthora infestans in progeny of dihaploid potato parents

Phytophthora infestans (Mont.) de Bary is the most important fungal pathogen of the potato (Solanum tuberosum). The introduction of major genes for resistance from the wild species S. demissum into potato cultivars is the earliest example of breeding for resistance using wild germplasm in this crop. Eleven resistance alleles (R genes) are known, differing in the recognition of corresponding avirulence alleles of the fungus. The number of R loci, their positions on the genetic map and the allelic relationships between different R variants are not known, except that the R1 locus has been mapped to potato chromosome V The objective of this work was the further genetic analysis of different R alleles in potato. Tetraploid potato cultivars carrying R alleles were reduced to the diploid level by inducing haploid parthenogenetic development of 2n female gametes. Of the 157 isolated primary dihaploids, 7 set seeds and carried the resistance alleles R1, R3 and R10 either individually or in combinations. Independent segregation of the dominant R1 and R3 alleles was demonstrated in two F₁ populations of crosses among a dihaploid clone carrying R1 plus R3 and susceptible pollinators. Distorted segregation in favour of susceptibility was found for the R3 allele in 15 of 18 F₁ populations analysed, whereas the RI allele segregated with a 1:1 ratio as expected in five F₁ populations. The mode of inheritance of the R10 allele could not be deduced as only very few F₁ hybrids bearing R10 were obtained. Linkage analysis in two F₁ populations between R1, R3 and RFLP markers of known position on the potato RFLP maps confirmed the position of the R1 locus on chromosome V and localized the second locus, R3, to a distal position on chromdsome XI.     

Cross adaptation of potato plants to low temperatures and potato cyst nematode infestation

Effect of short-term (2 h a day) and long-term (6 days) exposure to low temperature (5°C) on cold tolerance was investigated in two cultivars of potato (Solanum tuberosum L.): resistant (Sudarynya) and susceptible (Nevskii) to potato cyst nematode (Globodera rostochiensis Woll.). The extent of their infestation and changes in the expression of the genes of resistance to nematode (H1 and Gro1-4) were also analyzed. In both cultivars, exposure to low temperature enhanced cold resistance of potato plants. Enhancing cold resistance of cv. Sudarynya induced by a short-term exposure to chilling did not affect the extent of nematode infestation, whereas in susceptible cv. Nevskii, the extent of infestation decreased by almost three times. The level of expression of H1 gene in the leaves of the susceptible cultivar rose almost twofold both after short-term and long exposure to chilling, while in the resistant cultivar, gene expression increased only after a short-term effect of cold. The level of Gro1-4 gene expression increased after both temperature treatments only in the resistant cv. Sudarynya. Thus, the expression of genes for potato resistance to nematode infestation became more active in the susceptible cultivar as regards the gene H1 and in the resistant cultivar, regarding the gene Gro1-4. In the nematode-susceptible cv. Nevskii, the level of infestation decreased and cold resistance increased, apparently indicating cross adaptation to two factors of different nature.     

Mapping and marker-assisted selection for a gene for extreme resistance to potato virus Y

The chromosomal location of the major gene Ry _adg controlling extreme resistance to potato virus Y (PVY) in Solanum tuberosum subsp. andigena was identified by RFLP analysis of a diploid potato population. A total of 64 tomato and potato RFLP markers were screened with the bulked segregant analysis (BSA) on segregants extremely resistant, hypersensitive or susceptible to PVY. Four markers TG508, GP125, CD17 and CT168 at the proximal end of chromosome XI showed close linkage with extremely resistant phenotypes. TG508 was identified as the closest marker linked with the Ry _adg locus with the maximum map distance estimated as 2.0 cM. The 4 markers linked with the Ry _adg locus were tested on independent tetraploid and diploid potato clones and were subsequently found useful for marker-assisted selection for plants containing Ry _adg . Received: 5 July 1996 / Accepted: 19 July 1996     

Comparative sequence analysis of the potato cyst nematode resistance locus <Emphasis Type="Italic">H1</Emphasis> reveals a major lack of co-linearity between three haplotypes in potato (<Emphasis Type="Italic">Solanum tuberosum</Emphasis> ssp.)

The H1 locus confers resistance to the potato cyst nematode Globodera rostochiensis pathotypes 1 and 4. It is positioned at the distal end of chromosome V of the diploid Solanum tuberosum genotype SH83-92-488 (SH) on an introgression segment derived from S. tuberosum ssp. andigena. Markers from a high-resolution genetic map of the H1 locus (Bakker et al. in Theor Appl Genet 109:146–152, 2004) were used to screen a BAC library to construct a physical map covering a 341-kb region of the resistant haplotype coming from SH. For comparison, physical maps were also generated of the two haplotypes from the diploid susceptible genotype RH89-039-16 (S. tuberosum ssp. tuberosum/S. phureja), spanning syntenic regions of 700 and 319 kb. Gene predictions on the genomic segments resulted in the identification of a large cluster consisting of variable numbers of the CC-NB-LRR type of R genes for each haplotype. Furthermore, the regions were interspersed with numerous transposable elements and genes coding for an extensin-like protein and an amino acid transporter. Comparative analysis revealed a major lack of gene order conservation in the sequences of the three closely related haplotypes. Our data provide insight in the evolutionary mechanisms shaping the H1 locus and will facilitate the map-based cloning of the H1 resistance gene.     

Mapping a new nematode resistance locus in Lycopersicon peruvianum

Accessions of the wild tomato species L. peruvianum were screened with a root-knot nematode population (557R) which infects tomato plants carrying the nematode resistance gene Mi. Several accessions were found to carry resistance to 557R. A L. peruvianum backcross population segregating for resistance to 557R was produced. The segregation ratio of resistant to susceptible plants suggested that a single, dominant gene was a major factor in the new resistance. This gene, which we have designated Mi-3, confers resistance against nematode strains that can infect plants carrying Mi. Mi-3, or a closely linked gene, also confers resistance to nematodes at 32°C, a temperature at which Mi is not effective. Bulked-segregant analysis with resistant and susceptible DNA pools was employed to identify RAPD markers linked to this gene. Five-hundred-and-twenty oligonucleotide primers were screened and two markers linked to the new resistance gene were identified. One of the linked markers (NR14) was mapped to chromosome 12 of tomato in an L. esculentum/L. pennellii mapping population. Linkage of NR14 and Mi-3 with RFLP markers known to map on the short arm of chromosome 12 was confirmed by Southern analysis in the population segregating for Mi-3. We have positioned Mi-3 near RFLP marker TG180 which maps to the telomeric region of the short arm of chromosome 12 in tomato.     

QTL analysis of potato tuberization

Quantitative trait loci (QTLs) affecting tuberization were detected in reciprocal backcrosses between Solanum tuberosum and S. berthaultii. Linkage analyses were performed between traits and RFLP alleles segregating from both the hybrid and the recurrent parent using a set of framework markers from the potato map. Eleven distinct loci on seven chromosomes were associated with variation in tuberization. Most of the loci had small effects, but a QTL explaining 27% of the variance was found on chromosome 5. More QTLs were detected while following alleles segregating from the recurrent S. tuberosum parent used to make the backcross than were detected by following alleles segregating from the hybrid parent. More than half of the alleles favoring tuberization were at least partly dominant. Tuberization was favored by an allele from S. berthaultii at 3 of the 5 QTLs detected by segregation from the hybrid parent. The additive effects of the QTLs for tuberization explained up to 53% of the phenotypic variance, and inclusion of epistatic effects increased this figure to 60%. The most common form of epistasis was that in which presence of an allele at each of 2 loci favoring tuberization was no more effective than the presence of a favorable allele at 1 of the 2 loci. The QTLs detected for tuberization traits are discussed in relationship to those previously detected for trichome-mediated insect resistance derived from the unadapted wild species.Paper number 54 of the Department of Fruit and Vegetable Science, Cornell University     

The R1 gene conferring race-specific resistance to Phytophthora infestans in potato is located on potato chromosome V.

Summary Late blight in potato is caused by the fungusPhytophthora infestans and can inflict severe damage on the potato crop. Resistance toP. infestans is either based on major dominantR genes conferring vertical, race-specific resistance or on minor genes inducing horizontal, unspecific resistance. A dihaploid potato line was identified which carried theR1 gene, conferring vertical resistance to allP. infestans races, with the exception of those homozygous for the recessive virulence allele of the locusV1. The F₁ progeny of a cross between this resistant parent P(R1) and P(r), a line susceptible to all races, was analysed for segregation ofR1 and of restriction fragment length polymorphism (RFLP) markers distributed on the potato RFLP map comprising more than 300 loci. TheR1 locus was mapped to chromosome V in the interval between RFLP markers GP21 and GP179. The map position ofR1 was found to be very similar to the one ofRx2, a dominant locus inducing extreme resistance to potato virus X.     

Resistance to late blight in Solanum bulbocastanum is mapped to chromosome 8

Somatic hybrids between potato and Solanum bulbocastanum, a wild diploid (2n=2x=24) Mexican species, are highly resistant to late blight, caused by Phytophthora infestans. Both randomly amplified polymorphic DNA (RAPD) and restriction fragment length polymorphism (RFLP) markers that are closely linked to the resistance have been noted by analysis of three different backcross-2 populations derived from two different somatic hybrids. With reference to previously published potato and tomato maps, resistance appears to be on the long arm of chromosome 8 and is flanked by RFLP markers CP53 and CT64. In a population of BC₂ plants derived from a cross between the BC₁ line J10lK6 [(S. tuberosum PI 203900+S. bulbocastanum PI 243510) ×Katahdin)]×Atlantic, late blight resistance cosegregated with RFLP marker CT88 and RAPD marker OPG02–625. Received: 26 November 1999 / Accepted: 22 December 1999     

Characterization of the Aspergillus niger pelB gene: structure and regulation of expression

Summary Nearly isogenic lines (NILs) of rice (Oryza sativa) differing at a locus conferring resistance to the pathogen Xanthomonas oryzae pv. oryzae were surveyed with 123 DNA markers and 985 random primers using restriction fragment length plymorphism (RFLP) and random amplified polymorphic DNA (RAPD) analysis. One chromosome 11 marker (RG103) detected polymorphism between the NILs that cosegregated with Xa21. All other chromosome 11 DNA markers tested were monomorphic between the NILs, localizing the Xa21 introgressed region to an 8.3 cM interval on chromosome 11. Furthermore, we identified two polymerase chain reaction (PCR) products (RAPD2148 and RAPD818) that detected polymorphisms between the NILs. Genomic sequences hybridizing with RAPD818, RAPD248 and RG103 were duplicated specifically in the Xa21 NIL. All three markers cosegregated with the resistance locus, Xa21, in a F₂ population of 386 progeny. Based on the frequency with which we recovered polymorphic Xa21-linked markers, we estimated the physical size of the introgressed region to be approximately 800 kb. This estimation was supported by physical mapping (using pulsed field gel electrophoresis) of the sequences hybridizing with the three Xa21-linked DNA markers. The results showed that the three Xa21-linked markers are physically close to each other, with one copy of the RAPD818 sequences located within 60 kb of RAPD248 and the other copy within 270 kb of RG103. None of the enzymes tested generated a DNA fragment that hybridized with all three of the markers indicating that the introgressed region containing the resistance locus Xa21 is probably larger than 270 kb.     

Allelic state of the molecular marker for golden nematode (Globodera rostochiensis) resistance gene H1 among Ukrainian and world potato (Solanum tuberosum ssp. tuberosum) cultivars

The purpose of this study was to determine the allelic state of the resistance gene H1 against the Ro1 and Ro4 pathotypes of the golden potato cyst nematode (Globodera rostochiensis) among Ukrainian and world potato (Solanum tuberosum ssp. tuberosum) cultivars. The allelic state of the TG689 marker was determined by PCR with DNA samples isolated from potato tubers and primers, one pair of which flanked the allele-specific region and the other one was used to control the DNA quality. Among 77 potato cultivars analyzed, the allele of the marker associated with the H1-type resistance was found in 74% of Ukrainian and 90% of foreign cultivars, although some of them proved to be susceptible to the potato cyst nematode in the field. The obtained data confirm the presence of H1 resistance against golden nematode pathotypes Ro1 and Ro4 among Ukrainian potato cultivars and the efficiency of the used marker within the accuracy that has been declared by its authors.