An ultradense genetic linkage map with .10,000 AFLP loci was constructed from a heterozygous diploid potato population. To our knowledge, this is the densest meiotic recombination map ever constructed. A fast marker-ordering algorithm was used, based on the minimization of the total number of recombination events within a given marker order in combination with genotyping error-detection software. This resulted in ''skeleton bin maps,'' which can be viewed as the most parsimonious marker order. The unit of distance is not expressed in centimorgans but in ''bins.'' A bin is a position on the genetic map with a unique segregation pattern that is separated from adjacent bins by a single recombination event. Putative centromeres were identified by a strong clustering of markers, probably due to cold spots for recombination. Conversely, recombination hot spots resulted in large intervals of up to 15 cM without markers. The current level of marker saturation suggests that marker density is proportional to physical distance and independent of recombination frequency. Most chromatids (92%) recombined once or never, suggesting strong chiasma interference. Absolute chiasma interference within a chromosome arm could not be demonstrated. Two examples of contig construction and map-based cloning have demonstrated that the marker spacing was in accordance with the expected physical distance: approximately one marker per BAC length. Currently, the markers are used for genetic anchoring of a physical map of potato to deliver a sequence-ready minimal tiling path of BAC contigs of specific chromosomal regions for the potato genome sequencing consortium (http:/ /www.potatogenome.net).

Many plant and animal immune receptors have a modular nucleotide-binding-leucine-rich repeat (NB-LRR) architecture in which a nucleotide-binding switch domain, NB-ARC, is tethered to a LRR sensor domain. The cooperation between the switch and sensor domains, which regulates the activation of these proteins, is poorly understood. Here, we report structural determinants governing the interaction between the NB-ARC and LRR in the highly homologous plant immune receptors Gpa2 and Rx1, which recognize the potato cyst nematode Globodera pallida and Potato virus X, respectively. Systematic shuffling of polymorphic sites between Gpa2 and Rx1 showed that a minimal region in the ARC2 and N-terminal repeats of the LRR domain coordinate the activation state of the protein. We identified two closely spaced amino acid residues in this region of the ARC2 (positions 401 and 403) that distinguish between autoactivation and effector-triggered activation. Furthermore, a highly acidic loop region in the ARC2 domain and basic patches in the N-terminal end of the LRR domain were demonstrated to be required for the physical interaction between the ARC2 and LRR. The NB-ARC and LRR domains dissociate upon effector-dependent activation, and the complementary-charged regions are predicted to mediate a fast reassociation, enabling multiple rounds of activation. Finally, we present a mechanistic model showing how the ARC2, NB, and N-terminal half of the LRR form a clamp, which regulates the dissociation and reassociation of the switch and sensor domains in NB-LRR proteins.Resistance (R) proteins play a central role in the recognition-based immune system of plants. Unlike vertebrates, plants lack an adaptive immune system with highly specialized immune cells. Instead, they rely on an innate immune system in which each cell is autonomous. Two types of immune receptors can be distinguished in plants, pathogen-associated molecular patterns recognition receptors that detect conserved molecular patterns in plant pathogens and intracellular R proteins that recognize specific effectors employed by pathogens as modifiers of host metabolism or defense mechanisms (Jones and Dangl, 2006). Effector-triggered activation of R proteins leads to an array of protective responses, often culminating in programmed cell death at the site of infection (Greenberg and Yao, 2004), thereby preventing further ingress of the pathogen. Pathogens have evolved mechanisms to evade recognition by R proteins and to regain their virulence (Dodds and Rathjen, 2010). This continuous coevolutionary process between host and pathogen has resulted in a reservoir of highly diverse R proteins in plants, enabling them to counteract a wide range of pathogens and pests.The most common class of R proteins consists of nucleotide-binding (NB)-leucine-rich repeat (LRR) proteins with a tripartite domain architecture, which roughly corresponds to an N-terminal response domain (a coiled coil [CC] or Toll/Interleukin-1 receptor [TIR] domain) involved in downstream signaling, a central molecula...

SUMMARY Some plant endoparasitic nematodes are biotrophic and induce remarkable changes in their hosts in order to ensure a continuous supply of food. Proteins secreted from oesophageal gland cells have been implicated in this pathogenic process. A potentially secreted chorismate mutase has been isolated from the potato cyst nematode Globodera pallida. The gene encoding this protein is expressed in the subventral oesophageal gland cells of the nematode, and the mRNA derived from this gene is only present in the early parasitic stages. Sequence analysis of this gene shows that, like other genes involved in the host-parasite interaction of plant parasitic nematodes, it is likely to have been acquired by horizontal gene transfer from bacteria. The presence of a signal peptide in the deduced amino acid sequence of the G. pallida chorismate mutase and its expression in the subventral oesophageal gland cells suggest that it is secreted from the nematode, pointing to a role for the protein in the host-parasite interaction. The shikimate pathway, of which chorismate mutase is normally a part, is not found in animals but is present in plants and bacteria. In plants it gives rise to a variety of compounds which are important in amino acid synthesis and defence signalling pathways, as well as auxins, which have been implicated in the early development of nematode feeding sites. The potential roles of a nematode chorismate mutase are discussed.

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Like all plants, potato has evolved a surveillance system consisting of a large array of genes encoding for immune receptors that confer resistance to pathogens and pests. The majority of these so-called resistance or R proteins belong to the super-family that harbour a nucleotide binding and a leucine-rich-repeat domain (NB-LRR). Here, sequence information of the conserved NB domain was used to investigate the genome-wide genetic distribution of the NB-LRR resistance gene loci in potato. We analysed the sequences of 288 unique BAC clones selected using filter hybridisation screening of a BAC library of the diploid potato clone RH89-039-16 (S. tuberosum ssp. tuberosum) and a physical map of this BAC library. This resulted in the identification of 738 partial and full-length NB-LRR sequences. Based on homology of these sequences with known resistance genes, 280 and 448 sequences were classified as TIR-NB-LRR (TNL) and CC-NB-LRR (CNL) sequences, respectively. Genetic mapping revealed the presence of 15 TNL and 32 CNL loci. Thirty-six are novel, while three TNL loci and eight CNL loci are syntenic with previously identified functional resistance genes. The genetic map was complemented with 68 universal CAPS markers and 82 disease resistance trait loci described in literature, providing an excellent template for genetic studies and applied research in potato.Electronic supplementary materialThe online version of this article (doi:10.1007/s00122-011-1602-z) contains supplementary material, which is available to authorized users.

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.Electronic supplementary materialThe online version of this article (doi:10.1007/s00122-010-1472-9) contains supplementary material, which is available to authorized users.

The resistance gene H1 confers resistance to the potato cyst nematode Globodera rostochiensis and is located at the distal end of the long arm of chromosome V of potato. For marker enrichment of the H1 locus, a bulked segregant analysis (BSA) was carried out using 704 AFLP primer combinations. A second source of markers tightly linked to H1 is the ultra-high-density (UHD) genetic map of the potato cross SH x RH. This map has been produced with 387 AFLP primer combinations and consists of 10,365 AFLP markers in 1,118 bins (http://www.dpw.wageningen-ur.nl/uhd/). Comparing these two methods revealed that BSA resulted in one marker/cM and the UHD map in four markers/cM in the H1 interval. Subsequently, a high-resolution genetic map of the H1 locus has been developed using a segregating F(1) SH x RH population consisting of 1,209 genotypes. Two PCR-based markers were designed at either side of the H1 gene to screen the 1,209 genotypes for recombination events. In the high-resolution genetic map, two of the four co-segregating AFLP markers could be separated from the H1 gene. Marker EM1 is located at a distance of 0.2 cM, and marker EM14 is located at a distance of 0.8 cM. The other two co-segregating markers CM1 (in coupling) and EM15 (in repulsion) could not be separated from the H1 gene.

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