D. R. Lynch

Tomato Ve disease resistance genes encode cell surface-like receptors

In tomato, Ve is implicated in race-specific resistance to infection by Verticillium species causing crop disease. Characterization of the Ve locus involved positional cloning and isolation of two closely linked inverted genes. Expression of individual Ve genes in susceptible potato plants conferred resistance to an aggressive race 1 isolate of Verticillium albo-atrum. The deduced primary structure of Ve1 and Ve2 included a hydrophobic N-terminal signal peptide, leucine-rich repeats containing 28 or 35 potential glycosylation sites, a hydrophobic membrane-spanning domain, and a C-terminal domain with the mammalian E/DXXXLφ or YXXφ endocytosis signals (φ is an amino acid with a hydrophobic side chain). A leucine zipper-like sequence occurs in the hydrophobic N-terminal signal peptide of Ve1 and a Pro-Glu-Ser-Thr (PEST)-like sequence resides in the C-terminal domain of Ve2. These structures suggest that the Ve genes encode a class of cell-surface glycoproteins with receptor-mediated endocytosis-like signals and leucine zipper or PEST sequences.

Genetic diversity of potato determined by random amplified polymorphic DNA analysis.

SummaryThe RAPD procedure was used to establish genetic diversity of 28 potato genotypes including siblings and genotypes with no immediate relationship. In addition amplified DNA from three parents and Solanum chacoense were compared with that from six progeny to determine the genetic relationships. Amplification of genomic DNA from the 28 genotypes using PCR and 12 decamer primers yielded 158 amplified DNA fragments, ranging in size from 490 to 3200 bp. A total of 128 unique RAPD fragments were observed among the 28 potato genotypes. Similarity measures and principal coordinate analysis generally reflected the expected trends in relationships of the full and half-sib potato genotypes. However there were important exceptions to this general trend and it appears that related varieties can be as genetically different as varieties with no immediate relationship. The data suggest that RAPD analysis used in conjunction with pedigree information can provide a superior measure of genetic divergence than analysis based solely on pedigree information.

The effect of moisture stress at three growth stages on the yield, components of yield and processing quality of eight potato varieties

A study was conducted over a three year period in southern Alberta to establish the yield and quality responses of eight potato varieties-Atlantic, Conestoga, Norchip, Niska, Shepody, Ranger Russet (Idaho clone), Ranger Russet (Amisk clone) and Russet Burbank to a single transient moisture stress (-80 kPa) at three growth stages in crop development (early, midseason and late). Early and midseason moisture stress had the greatest negative impact (P<0.05) on tuber yield. Of the eight varieties, Atlantic and Conestoga appear to be particularly sensitive to stress at these two growth stages. Midseason stress also appeared to reduce specific gravity. A year x stress interaction (P<0.05) for fry color suggests that the influence of moisture stress on fry color can be modified by other environmental factors. The results of the study demonstrate the importance of maintaining adequate soil moisture at all stages during crop development.CompendioSe condujo un estudio en un periodo de tres años al sur de Alberta, para establecer la respuesta en rendimiento y calidad de ocho variedades [Atlantic, Conestoga, Norchip, Niska, Shepody, Ranger Russet (clon de Idaho), Ranger Russet (clon de Amisk) y Russet Burbank], a un estrés temporal a la humedad (-80 kPa) en tres estados de crecimiento durante el desarrollo del cultivo (temprano, a media temporada y tardíamente). El estrés al comienzo y a mediados de la temporada tuvo el mayor impacto negativo (P<0.05) sobre el rendimiento en tubérculos. De las ocho variedades, Atlantic y Conestoga parecen ser particularmente sensibles al estrés en estos dos estados de crecimiento. El estrés, a mitad de temporada, parece que también disminuyó la gravedad específica. Una interacción año x estrés (P<0.05), para color de fritura, sugiere que la influencia del estres sobre éste puede ser modifïcada por otros factores ambientales. Los resultados del estudio demuestran la importancia de mantener una humedad adecuada en el suelo en todos los estados durante el desarrollo del cultivo.

Characterization ofSolanum tuberosum simple sequence repeats and application to potato culiwar identification

With the continued introduction of new potato cultivars, accurate identification is becoming difficult but is essential for maintaining cultivar integrity and Plant Breeders’ Rights. Hypervariable DNA sequences, referred to as simple sequence repeats (SSRs) or microsatellites, have been reported to be an excellent source of genetic markers. To determine the abundance, distribution, and composition of SSRs withinSolanium tuberosum, 252 sequences were searched for tetranucleotide and smaller SSRs with a minimum length of 20 nucleotides and a maximum discrepancy of two nucleotides. In total, 40 unique SSRs were observed in the 252S. tuberosum sequences examined and occurred at a frequency of one SSR every 8.1 kb. To assess the ability of site-specific amplified SSRs to identify potato cultivars, a simple (TCAC)m and compound (TCAC)m • (CTT)n SSR 5’ to the starch synthase gene and a compound (C)p • (CT)q • (AT)r • (G)s SSR 5’ to the sequence encoding mature proteinase inhibitor I, were examined and shown to produce unique DNA profiles for 73 of 95 tetraploid cultivars. In total, 24 alleles were observed at these loci and the accurately sized amplified DNA products can be used to establish a database for cultivar identification. Site-specific amplified alleles were somatically stable and have been conserved in clonal variants of Russet Burbank independently maintained for almost seven decades, a characteristic essential for cultivar identification. As genetic markers, the abundant, informative, and easily examined site-specific amplified alleles of SSRs are ideal for quickly and accurately determining cultivar identity of S.tuberosum ssp.tuberosum.

Resistance to thiabendazole and thiophanate-methyl in canadian isolates ofFusarium sambucinum andHelminthosporium solani

Resistance to both thiabendazole (TBZ) and thiophanate-methyl (TPM) was observed in Canadian isolates ofFusarium sambucinum andHelminthosporium solani recovered from diseased potato tubers. Resistance was not found in isolates ofFusarium avenaceum, Fusarium culmorum, Fusarium equiseti andFusarium solani. A high level of resistance to both TBZ (EC50 180 to ≫500 mg/l) and TPM (EC50 ≫500 mg/l) was observed forH. solani isolates, whereasF. sambucinum exhibited a relatively low level of resistance to TBZ (EC50 34 to 71 mg/l) and a high level of resistance to TPM (growth uninhibited by 2,500 mg/l). The incidence ofF. sambucinum resistant to the benzimidazoles was lower (60%) than that observed forH. solani (95%)CompendioEn Canadá, se observó resistencia tanto al Tiabendazole como al MetilTiofanato, en aislamientos de Fusarium sambucinum y Helminthosporium solani obtenidos de tubérculos de papa enfermos. No se encontró resistencia en aislamientos de Fusarium avenaceum, Fusarium culmorum, Fusarium equiseti y Fusarium solani. Se observó un alto nivel de resistencia tanto al TBZ (EC50 180 a ≫500 mg/dm3) como al TPM (EC50 ≫500 mg/dm3) para los aislamientos de H. solani, mientras que F. sambucinum mostró un nivel relativamente bajo de resistencia al TBZ (EC50 34 a 71 mg/dm3) y un alto nivel de resistencia al TPM (su crecimiento no fue inhibido por 2 500 mg/dm3). La incidencia de F. sambucinum resistente a los bencimidazoles fue menor (60%) que la observada para H. solani (95%)

Identification of potato cultivars and clonal variants by random amplified polymorphic DNA analysis

Random Amplified Polymorphic DNAs (RAPDs) were used to distinguish commercial potato cultivars and clonal variants of cultivars. Primer 131, one of four primers used, distinguished 30 of the 36 cultivars tested. All 36 commercial cultivars were distinguished using only two primers (131 and 184). The RAPD pattern of 20 unidentified potato cultivars was compared with known patterns of 36 cultivars. Each one of the 20 cultivars was correctly identified. Particular primers appear to produce greater numbers of both amplified DNA fragments and polymorphisms, and are therefore suited to RAPD identification of potato cultivars. Polymorphism was obtained between Russet Burbank Idaho D and Russet Burbank White Skin with primer 251 and between Viking and Purple Viking with primer 380. However, polymorphism was not observed between Norgold, Norland, Sebago and Superior clones using only 20 primers. The RAPD technique is much more likely to detect polymorphism, regardless of tissue or environmental factors, than isozyme analysis and is easier, less costly and faster than the RFLP procedure. Thus, RAPD analysis represents a highly useful method of distinguishing and identifying potato cultivars and clonal variants of cultivars.CompendioSe utilizaron DNAs Polimórficos Amplificados al Azar (RAPDs) para identificar cultivares comerciales de papa y variantes clonales de cultivares. El Imprimador 131, uno de cuatro imprimadores utilizados, identificó 30 de los 36 cultivares estudiados. Todos los 36 cultivares comerciales fueron identificados utilizando solamente dos imprimadores (131 y 184). El modelo RAPD de 20 cultivares no identificados fue comparado con modelos conocidos de 36 cultivares. Cada uno de los 20 cultivares fue correctamente identificado. Imprimadores específicos parecen producir números mayores tanto de fragmentos como de polimorfismos amplificados de DNA y son por lo tanto apropiados para la identificatión de cultivares de papa utilizando el RAPD. Se obtuvo polimorfismo entre Russet Burbank Idaho D y Russet Burbank White Skin con el imprimador 251 y entre Viking y Purple Viking con el imprimador 380. Sin embargo, utilizando solamente 20 imprimadores, no se observaron polimorfismos entre los clones de Norgold, Norland, Sebago y Superior. La técnica RAPD es más apropiada para detectar polismorfismos, independientemente del tejido o de los factores del ambiente, que el análisis de isoenzimas y es más fácil, menos costosa y más rápida que el procedimiento RFLP (polimorfismo de restricción de la longitud de los fragmentes). Por lo tanto, el análisis RAPD representa un método muy útil para distinguir e identificar cultivares de papa y variantes clonales de los mismos.

Isolation of the β-tubulin gene and characterization of thiabendazole resistance in Gibberella pulicaris

Gibberella pulicaris (anamorph: Fusarium sambucinum), is a common potato pathogen that produces trichothecene mycotoxins and reduces plant yield and tuber quality. Field isolates have been recovered from western Canada that are resistant to thiabendazole, a benzimidazole fungicide registered in many countries to control dry rot in storage. To characterize this resistance, homokaryons were derived from single ascospores that were produced in vitro from crosses of thiabendazole-resistant G. pulicaris field isolates with sensitive isolates of an opposite mating type. The β-tubulin gene was amplified from these homokaryons using degenerate oligonucleotides synthesized to conserved sequences of the β-tubulin gene derived from related fungi. An open reading frame containing three putative introns and a deduced amino acid sequence of 446 amino acids exhibited a high level of homology to the β-tubulin gene of other fungi. Although nucleotide and amino acid differences were observed in the β-tubulin gene, none were linked to the thiabendazole resistance. Linkage analysis confirmed that the isolated β-tubulin gene is single copy and was not linked to thiabendazole resistance in G. pulicaris.

Identification of late blight, Colorado potato beetle, and blackleg resistance in three Mexican and two South American wild 2x (1EBN) Solanum species

Wild potatoes are important sources of genes for resistance to disease and insect pests. A collection of wild Mexican and South AmericanSolarium species from the US potato Genebank was evaluated under laboratory and/or field conditions for their reaction to late blight (Phytophthora infestans), Colorado potato beetle (CPB,Leptinotarsa decemlineata Say), and blackleg (Erwinia carotovora subsp.atroseptica (van Hall) Dye) in order to identify individual genotypes with multiple resistance genes. Late blight inoculations using aggressive isolates (US-8/A2 and US-11/A1 mating types) of P.infestans revealed a wide range of variation for resistance between and within the accessions of the wild species tested. For late blight, susceptible as well as moderately to highly resistant genotypes were observed in all the species tested. However, at least one accession from the three Mexican and one South American wild diploid species tested showed a relatively uniform high level of resistance toP. infestans. These includedS. bulbocastanum, S. pinnatisectum, S. cardiophyllum, andS. circaeifolium. Two accessions from South American speciesS. commersonii were highly susceptible to late blight. For the Colorado potato beetle test, only one species,S. pinnatisectum appeared uniformly resistant to CPB under field conditions. Results of screening for blackleg resistance showed that there were major differences between genotypes in the wild species. Accessions ofS. circaeifolium PI 498119 andS. bulbocastanum PI 243504 were identified as having significantly higher blackleg resistance than cultivated potato and the other wild species tested. However, genotypes from these two accessions were more susceptible to late blight and CPB. Characterization of theP. infestans isolate P1801C.16 used for late blight evaluation and multi-locus isolate tests using US-8/A2 and US-11/A1 races revealed that the resistance inS. pinnatisectum genotypes tested corresponded to a race-non-specific genetic system, which was different from any existing R genes.Solanum pin-natisectum genotypes with both high levels of late blight and CPB resistance as well as blackleg resistance genotypes identified in the present study represent a diverse gene pool that may be useful for development of new potato cultivars with multiple disease and insect resistance. The potential utilization of these valuable sources for improvement of cultivated potato is discussed.ResumenLas papas silvestres son la fuente importante de genes de resistencia a las enfermedades y piagas. Una colección de especies silvestres mexicanas y sudamericanas deSolanum del banco de genes de papa de EUA fue evaluada bajo condiciones de laboratorio y campo para reacción al tizón tardio (Phytophthora infestans), escarabajo Colorado de la papa (CPB,Leptinotarsa decemlineata Say) y pierna negra (Erwinia carotovora subsp.atroseptica (van Hall) Dye) con el objeto de identificar genotipos individuales con genes multiples de resistencia. Inoculaciones hechas usando aislamientos agresivos de P.infestans (tipos de apareamiento US-8/A2 y US-11/A1) revelaron un amplio rango de variación para resistencia entre y dentro de las accesiones de las especies silvestres probadas. Para tizón tardío se observaron tanto genotipos susceptibles como moderados a altamente resistentes en todas las especies probadas. Sin embargo, por lo menos una accesión de las especies silvestres diploides de las tres mexicanas y una sudamericana mostraron un alto nivel de resistencia relativamente uniforme aP. infestans. Estos incluyeronS. bulbocastanum, S. pinnatisectum, S. cardiophyllum yS. circaeifolium. Dos accesiones de la especie sudamericanaS. commersonii fueron altamente susceptible al tizón tardío. Solamente la especieS. pinnatisectum mostró resistencia uniforme al escarabajo Colorado bajo condiciones de campo. Los resultados del tamizado para resistencia a la pierna negra demostraron la existencia de diferencias mayores en las especies silvestres. Las accesiones deS. circaeifolium PI 498119 yS. bulbocas-tanum PI 243504 se identificaron como poseedoras de una resistencia significativamente alta a la pierna negra en comparacion con las especies cultivadas de papa, al igual que otras especies silvestres probadas. Sin embargo, los genotipos de estas dos accesiones fueron más susceptibles al tizón tardio y al escarabajo Colorado. La caracterización del aislamiento PI801C.16 usado para la evaluacién de tizón tardio y la prueba de aislamiento multi-locus utilizando las razas US-8/A2 y US-11/A1 revelaron que la resistencia en los genotipos probados deS. pinnatisectum, corresponden a una raza no específica del sistema genético, la cual era diferente de cualquiera de los genes R existentes. Los genotiposS. pinnatisectum con altos niveles de resistencia a tizón tardío y al escarabajo Colorado, lo mismo que los genotipos de resistencia a la pierna negra, identificados en el presente estudio representan un conjunto de genes que pueden ser utiles para el desarrollo de cultivares nuevos de papa con multiple resistencia a enfermedades y plagas. Se discute la utilizatión potencial de estas valiosas fuentes de resistencia para el mejoramiento de papa cultivada.

Field resistance to the potato leafroll luteovirus in transgenic and somaclone potato plants reduces tuber disease symptoms

Potato plants transformed with one of two potato leafroll luteovirus (PLRV) coat protein cDNA sequences and somaclonal variants recovered from the transformation process were examined to determine agronomic characteristics, incidence and level of resistance to PLRV, and the severity of disease symptoms. In total, 97 genetically modified lines of the potato cultivar Russet Burbank were initially screened in a greenhouse and selected lines subsequently examined in three independent field trials using a multiple year randomized block design. Abnormal plant phenotypes were observed in all lines and included reduced plant height, chlorotic leaves, delayed flowering, abnormal flower structure, and low tuber yields. Mean titres of PLRV in the field experiments, determined by enzyme-linked immunosorbent assay, were reduced by 32% to 71% and the incidence of elevated infection (>0.10 ng of PLRV per mg of leaf tissue) was reduced by 24% to 45% in the modified lines. Resistance to PLRV was sustained with similar virus titres in primary and secondary PLRV infections and was observed through 8 years of vegetative propagation and 5 years of field evaluation. Primary and secondary symptoms of PLRV infection were observed in the modified and wild type plants. However, the incidence and severity of internal tuber necrosis was significantly reduced in each of the modified lines. This is the first report characterising field performance of PLRV resistant plants transformed with the PLRV coat protein cDNA or derived through somaclonal variation.

Rhizoctonia disease of potatoes (Rhizoctonia solani): Fungicidal efficacy and cultivar susceptibility

Rhizoctonia stem canker and black scurf is an economically important disease of potatoes in Alberta and around the world. It reduces the quality and yield of potatoes and has become an important impediment for export of seed potatoes, especially to Mexico. Seed treatment using fungicides, presently registered in Canada, are not effective in controlling the disease to growers’ satisfaction. Field experiments conducted to evaluate the efficacy of various fungicides showed that captan (Captan), iprodione (Rovral), mancozeb (Tuberseal), and fludioxonil (Maxim) provide effective control of the disease. Of these fungicides, fludioxonil consistently provided superior protection. Based on the results of this study, Maxim was registered in 2000 for control of this disease in Canada. Potato cultivars/breeding selections showed a range of susceptibility reactions toR. solani but none of these was completely resistant to the disease. The information on comparative susceptibility of potato cultivars will help the growers make informed decisions regarding the management of this disease. Of six plant species (Beta vulgaris,Brassica compestris, Hordeum vulgare, Pisum sativum,Triticum aestivum, Zea mays) most commonly used in rotation with potatoes in Alberta, none was infected by potato isolates ofR. solani. The results suggested that use of these plant species as rotation crops with potatoes in Alberta will help in reducing the development of rhizoctonia disease in potatoes.ResumenEl cancro del tallo y costra negra causado por la rizoctonia es una de las enfermedades económicamente más importantes de las papas en Alberta y otras partes del mundo. Esta enfermedad reduce la calidad y el rendimiento de los tubérculos y se ha convertido en una limitación importante para la exportación de semillas de papa especialmente a México. El tratamiento de las semillas usando fungicidas, de preferencia registrados en Canada, no cumple las expectativas de los agricultores sobre el control de la enfermedad. Los experimentos en campo realizados para evaluar la eficacia de diversos fungicidas muestran que el captan (Captan), iprodione (Rovral), mancozeb (Tuberseal) y fludioxonil (Maxim) proporcionan un control efectivo de la enfermedad. De todos ellos, el fludioxonil proporciona una protección superior. Basados en los resultados de esta estudio, en el 2000 se registró a Maxim para el control de esta enfermedad en Canadá. Los cultivares de papa/selecciones por mejoramiento mostraron una gama de reacciones de susceptibilidad a R. solani pero ninguno fue completamente resistente a la enfermedad. La información sobre susceptibilidad comparada de los cultivares de papa ayudará a los agricultores a tomar decisones sobre la base de una mejor información considerando el manejo de esta enfermedad. De las seis especies de plantas más comúnmente usadas en rotación con las papas en Alberta (Beta vulgaris, Brassica compestris, Hordeum vulgare, Pisum sativum, Triticum aestivum, Zea mays), ninguna fue infectada con aislamientos de R. solani. Esos resultados sugieren que el uso de esas especies de plantas como cultivos rotatorios con las papas en Alberta podrían ayudar a reducir el desarrollo de la rizoctonia.