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Background In flowering plants it has been shown that de novo genome assemblies of different species and genera show a significant drop in the proportion of alignable sequence. Within a plant species, however, it is assumed that different haplotypes of the same chromosome align well. In this paper we have compared three de novo assemblies of potato chromosome 5 and report on the sequence variation and the proportion of sequence that can be aligned. Results For the diploid potato clone RH89-039-16 (RH) we produced two linkage phase controlled and haplotype-specific assemblies of chromosome 5 based on BAC-by-BAC sequencing, which were aligned to each other and compared to the 52 Mb chromosome 5 reference sequence of the doubled monoploid clone DM 1–3 516 R44 (DM). We identified 17.0 Mb of non-redundant sequence scaffolds derived from euchromatic regions of RH and 38.4 Mb from the pericentromeric heterochromatin. For 32.7 Mb of the RH sequences the correct position and order on chromosome 5 was determined, using genetic markers, fluorescence in situ hybridisation and alignment to the DM reference genome. This ordered fraction of the RH sequences is situated in the euchromatic arms and in the heterochromatin borders. In the euchromatic regions, the sequence collinearity between the three chromosomal homologs is good, but interruption of collinearity occurs at nine gene clusters. Towards and into the heterochromatin borders, absence of collinearity due to structural variation was more extensive and was caused by hemizygous and poorly aligning regions of up to 450 kb in length. In the most central heterochromatin, a total of 22.7 Mb sequence from both RH haplotypes remained unordered. These RH sequences have very few syntenic regions and represent a non-alignable region between the RH and DM heterochromatin haplotypes of chromosome 5. Conclusions Our results show that among homologous potato chromosomes large regions are present with dramatic loss of sequence collinearity

This paper describes the progress that has been made since the draft genome sequence of potato has been obtained and the analyses that need to be done to make further progress. Although sequencing has become less expensive and read lengths have increased, making optimal use of the information obtained is still difficult, certainly in the tetraploid potato crop. Major challenges in potato genomics are standardized genome assembly and haplotype analysis. Sequencing methods need to be improved further to achieve precision breeding. With the current new generation sequencing technology, the focus in potato breeding will shift from phenotype improvement to genotype improvement. In this respect, it is essential to realize that different alleles of the same gene can lead to different phenotypes depending on the genetic background and that there is significant epistatic interaction between different alleles. Genome-wide association studies will gain statistical power when binary single nucleotide polymorphism (SNP) data can be replaced with multi-allelic haplotype data. Binary SNP can be distributed across the many different alleles per locus or may be haplotype-specific, and potentially tag specific alleles which clearly differ in their contribution to a certain trait value. Assembling reads from the same linkage phase proved to allow constructing sufficiently long haplotype tracts to ensure their uniqueness. Combining large phenotyping data sets with modern approaches to sequencing and haplotype analysis and proper software will allow the efficiency of potato breeding to increase.

Treatments of tubers with chemical disinfectants (70 % ethanol, 1 % sodium hypochlorite, 2 % copper sulphate, 5 % peracetic acid, 10 % hydrogen peroxide, 1 % MennoClean (benzoic acid), 5 % trisodium phosphate and 0.2 % caffeine) were evaluated for control of blackleg caused by ‘D. solani’. All disinfectants effectively killed bacteria in axenic cultures within 5 min and all, except hydrogen peroxide and trisodium phosphate, were able to kill ‘D. solani’ in spiked potato extracts. Treatments with all disinfectants except trisodium phosphate, reduced the pathogen populations on the periderm of tubers previously inoculated by dipping in a suspension of ‘D. solani’, when the disinfectant was applied for at least 15 min. However, in replicated greenhouse experiments, treatments with all disinfectants except hydrogen peroxide and caffeine resulted in phytotoxicity, reducing sprouting of tubers with 10 – 100 %. Sodium hypochlorite and MennoClean were selected for further studies. Treatments of inoculated tubers with these disinfectants prevented developing of soft rot symptoms on tubers when subjected to favourable rotting conditions. In replicated greenhouse experiments, inoculated tubers treated with sodium hypochlorite and MennoClean reduced the blackleg disease incidences from 50 % to 0 % and to 5 %, respectively, and symptomless infection of potato stems from 93 % to 0 % and to 5 %, respectively. A combination of both compounds was highly phytotoxic. The potential use of sodium hypochlorite and MennoClean in an integrated management strategy for potato blackleg is discussed., Treatments of tubers with chemical disinfectants (70 % ethanol, 1 % sodium hypochlorite, 2 % copper sulphate, 5 % peracetic acid, 10 % hydrogen peroxide, 1 % MennoClean (benzoic acid), 5 % trisodium phosphate and 0.2 % caffeine) were evaluated for control of blackleg caused by ‘D. solani’. All disinfectants effectively killed bacteria in axenic cultures within 5 min and all, except hydrogen peroxide and trisodium phosphate, were able to kill ‘D. solani’ in spiked potato extracts. Treatments with all disinfectants except trisodium phosphate, reduced the pathogen populations on the periderm of tubers previously inoculated by dipping in a suspension of ‘D. solani’, when the disinfectant was applied for at least 15 min. However, in replicated greenhouse experiments, treatments with all disinfectants except hydrogen peroxide and caffeine resulted in phytotoxicity, reducing sprouting of tubers with 10 – 100 %. Sodium hypochlorite and MennoClean were selected for further studies. Treatments of inoculated tubers with these disinfectants prevented developing of soft rot symptoms on tubers when subjected to favourable rotting conditions. In replicated greenhouse experiments, inoculated tubers treated with sodium hypochlorite and MennoClean reduced the blackleg disease incidences from 50 % to 0 % and to 5 %, respectively, and symptomless infection of potato stems from 93 % to 0 % and to 5 %, respectively. A combination of both compounds was highly phytotoxic. The potential use of sodium hypochlorite and MennoClean in an integrated management strategy for potato blackleg is discussed.

The genome of potato, a major global food crop, was recently sequenced. The work presented here details the integration of the potato reference genome (DM) with a new STS marker based linkage map and other physical and genetic maps of potato and the closely related species tomato. Primary anchoring of the DM genome assembly was accomplished using a diploid segregating population, which was genotyped with several types of molecular genetic markers to construct a new ~936 cM linkage map comprising 2,469 marker loci. In silico anchoring approaches employed genetic and physical maps from the diploid potato genotype RH and tomato. This combined approach has allowed 951 superscaffolds to be ordered into pseudomolecules corresponding to the 12 potato chromosomes. These pseudomolecules represent 674 Mb (~93%) of the 723 Mb genome assembly and 37,482 (~96%) of the 39,031 predicted genes. The superscaffold order and orientation within the pseudomolecules is closely collinear with independently constructed high density linkage maps. Comparisons between marker distribution and physical location reveal regions of greater and lesser recombination, as well as regions exhibiting significant segregation distortion. The work presented here has led to a greatly improved ordering of the potato reference genome superscaffolds into chromosomal 'pseudomolecules'.

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.

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

Potato (Solanum tuberosum L.) is the world’s most important non-grain food crop and is central to global food security. It is clonally propagated, highly heterozygous, autotetraploid, and suffers acute inbreeding depression. Here we use a homozygous doubled-monoploid potato clone to sequence and assemble 86% of the 844-megabase genome. We predict 39,031 protein-coding genes and present evidence for at least two genome duplication events indicative of a palaeopolyploid origin. As the first genome sequence of an asterid, the potato genome reveals 2,642 genes specific to this large angiosperm clade. We also sequenced a heterozygous diploid clone and show that gene presence/absence variants and other potentially deleterious mutations occur frequently and are a likely cause of inbreeding depression. Gene family expansion, tissue-specific expression and recruitment of genes to new pathways contributed to the evolution of tuber development. The potato genome sequence provides a platform for genetic improvement of this vital crop

Background Potato is the world's third most important food crop, yet cultivar improvement and genomic research in general remain difficult because of the heterozygous and tetraploid nature of its genome. The development of physical map resources that can facilitate genomic analyses in potato has so far been very limited. Here we present the methods of construction and the general statistics of the first two genome-wide BAC physical maps of potato, which were made from the heterozygous diploid clone RH89-039-16 (RH). Results First, a gel electrophoresis-based physical map was made by AFLP fingerprinting of 64478 BAC clones, which were aligned into 4150 contigs with an estimated total length of 1361 Mb. Screening of BAC pools, followed by the KeyMaps in silico anchoring procedure, identified 1725 AFLP markers in the physical map, and 1252 BAC contigs were anchored the ultradense potato genetic map. A second, sequence-tag-based physical map was constructed from 65919 whole genome profiling (WGP) BAC fingerprints and these were aligned into 3601 BAC contigs spanning 1396 Mb. The 39733 BAC clones that overlap between both physical maps provided anchors to 1127 contigs in the WGP physical map, and reduced the number of contigs to around 2800 in each map separately. Both physical maps were 1.64 times longer than the 850 Mb potato genome. Genome heterozygosity and incomplete merging of BAC contigs are two factors that can explain this map inflation. The contig information of both physical maps was united in a single table that describes hybrid potato physical map. Conclusions The AFLP physical map has already been used by the Potato Genome Sequencing Consortium for sequencing 10% of the heterozygous genome of clone RH on a BAC-by-BAC basis. By layering a new WGP physical map on top of the AFLP physical map, a genetically anchored genome-wide framework of 322434 sequence tags has been created. This reference framework can be used for anchoring and ordering of genomic sequenc

A cytogenetic map has been developed for diploid potato (Solanum tuberosum), in which the arms of the 12 potato bivalents can be identified in pachytene complements using multicolor fluorescence in situ hybridization (FISH) with a set of 60 genetically anchored bacterial artificial chromosome (BAC) clones from the RHPOTKEY BAC library. This diagnostic set of selected BACs (five per chromosome) hybridizes to euchromatic regions and corresponds to well-defined loci in the ultradense genetic map, and with these probes a new detailed and reliable pachytene karyotype could be established. Chromosome size has been estimated both from microscopic length measurements and from 4',6-diamidino-2-phenylindole fluorescence-based DNA content measurements. In both approaches, chromosome 1 is the largest (100–115 Mb) and chromosome 11 the smallest (49–53 Mb). Detailed measurements of mega-base-pair to micrometer ratios have been obtained for chromosome 5, with average values of 1.07 Mb/µm for euchromatin and 3.67 Mb/µm for heterochromatin. In addition, our FISH results helped to solve two discrepancies in the potato genetic map related to chromosomes 8 and 12. Finally, we discuss the significance of the potato cytogenetic map for extended FISH studies in potato and related Solanaceae, which will be especially beneficial for the potato genome-sequencing project

Potato is a member of the Solanaceae, a plant family that includes several other economically important species, such as tomato, eggplant, petunia, tobacco and pepper. The Potato Genome Sequencing Consortium (PGSC) aims to elucidate the complete genome sequence of potato, the third most important food crop in the world. The PGSC is a collaboration between 13 research groups from China, India, Poland, Russia, the Netherlands, Ireland, Argentina, Brazil, Chile, Peru, USA, New Zealand and the UK. The potato genome consists of 12 chromosomes and has a (haploid) length of approximately 840 million base pairs, making it a medium-sized plant genome. The sequencing project builds on a diploid potato genomic bacterial artificial chromosome (BAC) clone library of 78000 clones, which has been fingerprinted and aligned into ~7000 physical map contigs. In addition, the BAC-ends have been sequenced and are publicly available. Approximately 30000 BACs are anchored to the Ultra High Density genetic map of potato, composed of 10000 unique AFLPTM markers. From this integrated genetic-physical map, between 50 to 150 seed BACs have currently been identified for every chromosome. Fluorescent in situ hybridization experiments on selected BAC clones confirm these anchor points. The seed clones provide the starting point for a BAC-by-BAC sequencing strategy. This strategy is being complemented by whole genome shotgun sequencing approaches using both 454 GS FLX and Illumina GA2 instruments. Assembly and annotation of the sequence data will be performed using publicly available and tailor-made tools. The availability of the annotated data will help to characterize germplasm collections based on allelic variance and to assist potato breeders to more fully exploit the genetic potential of potato

In agriculture crop protection products (CPP) are used to protect the crop against pests, diseases and weeds. Several sprayings of CPP are done per crop per season. The used amount of CPP per spraying is generally based on a pre set CPP dose. Most often fields with variations in crop development are treated uniformly while spraying some patches in these fields would have been sufficient. In order to deal with the variations in crop development and site-specific variations in a field, a sensor-based spray technology was developed; SensiSpray. The system consists of sensors to detect crop variation and a spray system to automatically change spray volume. Electronics and software were developed to use the output signal of the sensor to adapt spray volume. For the variation of spray volume Lechler VarioSelect nozzle bodies were used fitted with four different low-drift venturi flat fan nozzles. On a 27 m working width boom sprayer seven sensors were placed on each boom section to control spray volume per boom section of 3-4.5m wide. Spray deposition measurements were performed to test the sprayer accuracy in adapting spray volume based on the reflection signal per section, using a fluorescent dye (Brilliant Sulpho Flavine) added to the spray liquid. The system is able to adapt spray volume within 1-2 m in the direction travelled. To demonstrate the potential use, the SensiSpray system was used in potato haulm killing spraying. The variation in reflection of the potato canopy was used to control the spray volume during desiccation spraying before harvest. MLHD PHK dosing algorithms for potato haulm killing herbicides were used to translate local greenness into a minimum effective dose. In the 2007 and 2008 season different potato fields were sprayed and a general use reduction in CPP for potato haulm killing was circa 50%, maintaining efficacy.

A 3-year case study was undertaken of how North American farmers use yield monitors for on-farm trials in farm management decision making. Case study methods were used because relatively few farmers quantitatively analyze yield monitor data. At this early research stage, insufficient farm management information about the data was available to ask the right questions in a large-scale survey. In addition to the formal case study of farmers experienced at using yield monitors to collect on-farm trial data, the study evaluated the effect of yield monitor data quality on farm decisions. Two levels of yield data quality included standard output where the default settings of farm-level mapping software were accepted and where filtering of the data was undertaken. Results indicated that yield data quality affects farm management decisions. In addition, farmers receiving a spatial analysis of their on-farm trial data tended to use split-field designs instead of replicated split-planter designs. They were also more confident in their decisions than before participation in the spatial analysis project, and made decisions more quickly. © 2008 Springer Science+Business Media, LLC.

In preparasitic second-stage juveniles (J2) of potato cyst nematode Globodera rostochiensis, six proteins with molecular masses of 30, 31a/b, 32, 39, and 49 kDa were recognized on Western blots by a monoclonal antibody (MGR48) specific for the subventral esophageal glands. All of these subventral gland proteins (svp's) focused in the basic range (pI 6.8 to 8.6) of an immobilized pH gradient. Western blotting showed that the svp's were present in preparasitic and parasitic J2 and not in later juvenile stages and adult females. Minor svp quantities also were observed in adult males. Immunogold labeling of preparasitic J2 showed that the svp's were localized in the rough endoplasmic reticulum and secretory granules of the subventral esophageal glands. Potato root diffusate triggered the secretion of svp's through the stylet, and 5-methoxy-N,N-dimethyltryptamine-hydrogen-oxalate had only a quantitative, additional effect. The forward flow of svp's through the metacorporal pump chamber was confirmed by the presence of svp's in the circular lumen of the esophagus (procorpus), as established by immunoelectron microscopy. Our data provide conclusive evidence that secretory proteins of the subventral glands of G. rostochiensis can be secreted through the stylet and support the hypothesis that the subventral esophageal glands play an important role in the early events of this nematode-plant interaction.

Secretory proteins from the dorsal and subventral oesophageal glands of potato cyst- nematodes (Globodera rostochiensis and G.pallida ) are considered to play an important role in the induction and exploitation of the specialized feeding cells (syncytia) that these parasites establish in the roots of their host plant. Identification of these secretory proteins will provide insight into the mechanisms involved in the formation of these syncytia, and is likely to aid in the development of novel forms of (artificial) host plant resistance to potato cyst-nematode infection. This thesis presents the data which were collected during a search for the oesophageal gland secretory proteins of G.rostochiensis.In the first part of the investigations (Chapters 2 to 5) the proteins of G.rostochiensis are examined using two-dimensional gel electrophoresis (2D-GE) followed by silver staining. Chapter 2 compares the protein patterns of two motile (2nd-stage juveniles (J2) and males) and two sedentary (4th-stage females and adult females) developmental stages using conventional (2D-GE) in large gels. This demonstrated very large differences in protein composition between these four developmental stages, and it was estimated that 74% of the polypeptides visible in these gels showed variations in their presence or abundance. The highest similarity index values were calculated for the protein patterns of both motile stages (0.674) and of both sedentary stages (0.767) with decreased values for all other pairwise comparisons.Chapter 3 introduces a miniature 2D-GE technique of which the high sensitivity is demonstrated by its ability to genotype single females of G. pallida . In Chapter 4 this mini 2D-GE technique is used to analyse the proteins of the complete range of post-embryonic developmental stages of G. rostochiensis. This analysis demonstrated large changes in protein pattern during the transformation of preparasitic J2 into sedentary J3 and during the metamorphosis of males into

Efficient and accurate diagnostic assays are essential for the design and evaluation of control measures of the potato cyst nematodesGlobodera rostochiensis andG. pallida by means of resistance. The hybridoma technology and the polymerase chain reaction (PCR) offer in potential various possibilities to design such diagnostic tests for routine purposes. We set out to devise a refined advisory system based on biochemical assays by using the following stepwise approach.

Two-dimensional polyacrylamide gel electrophoresis was used to examine the differences in total protein composition between two motile stages and two sedentary stages of the potato cyst-nematode, Globodera rostochiensis. Using a sensitive silver stain, 542 reproducible protein spots were distinguished. A list of these spots is presented, showing their apparent molecular weights, estimated isoelectric points, and occurrences in the different developmental stages. When the protein patterns were compared, 401 spots were found to change their presence or size in one or more of the four developmental stages. It is therefore estimated that during the post-embryonic development of G. rostochiensis, 74% of the polypeptides undergo modulation of their synthesis, or are affected by protein degradation or modification. In the motile stages several abundant proteins were present, which disappeared or decreased in concentration in the sedentary stages. Some of these proteins are presumably muscle proteins, and their modulation may illustrate the degeneration of body-wall musculature in the sedentary stages. It is concluded that the potato cyst-nematode has a very dynamic protein metabolism.