Joseph J. Coombs

Integration of Two Diploid Potato Linkage Maps with the Potato Genome Sequence

To facilitate genome-guided breeding in potato, we developed an 8303 Single Nucleotide Polymorphism (SNP) marker array using potato genome and transcriptome resources. To validate the Infinium 8303 Potato Array, we developed linkage maps from two diploid populations (DRH and D84) and compared these maps with the assembled potato genome sequence. Both populations used the doubled monoploid reference genotype DM1-3 516 R44 as the female parent but had different heterozygous diploid male parents (RH89-039-16 and 84SD22). Over 4,400 markers were mapped (1,960 in DRH and 2,454 in D84, 787 in common) resulting in map sizes of 965 (DRH) and 792 (D84) cM, covering 87% (DRH) and 88% (D84) of genome sequence length. Of the mapped markers, 33.5% were in candidate genes selected for the array, 4.5% were markers from existing genetic maps, and 61% were selected based on distribution across the genome. Markers with distorted segregation ratios occurred in blocks in both linkage maps, accounting for 4% (DRH) and 9% (D84) of mapped markers. Markers with distorted segregation ratios were unique to each population with blocks on chromosomes 9 and 12 in DRH and 3, 4, 6 and 8 in D84. Chromosome assignment of markers based on linkage mapping differed from sequence alignment with the Potato Genome Sequencing Consortium (PGSC) pseudomolecules for 1% of the mapped markers with some disconcordant markers attributable to paralogs. In total, 126 (DRH) and 226 (D84) mapped markers were not anchored to the pseudomolecules and provide new scaffold anchoring data to improve the potato genome assembly. The high degree of concordance between the linkage maps and the pseudomolecules demonstrates both the quality of the potato genome sequence and the functionality of the Infinium 8303 Potato Array. The broad genome coverage of the Infinium 8303 Potato Array compared to other marker sets will enable numerous downstream applications.

Retrospective View of North American Potato (Solanum tuberosum L.) Breeding in the 20th and 21st Centuries

Cultivated potato (Solanum tuberosum L.), a vegetatively propagated autotetraploid, has been bred for distinct market classes, including fresh market, pigmented, and processing varieties. Breeding efforts have relied on phenotypic selection of populations developed from intra- and intermarket class crosses and introgressions of wild and cultivated Solanum relatives. To retrospectively explore the effects of potato breeding at the genome level, we used 8303 single-nucleotide polymorphism markers to genotype a 250-line diversity panel composed of wild species, genetic stocks, and cultivated potato lines with release dates ranging from 1857 to 2011. Population structure analysis revealed four subpopulations within the panel, with cultivated potato lines grouping together and separate from wild species and genetic stocks. With pairwise kinship estimates clear separation between potato market classes was observed. Modern breeding efforts have scarcely changed the percentage of heterozygous loci or the frequency of homozygous, single-dose, and duplex loci on a genome level, despite concerted efforts by breeders. In contrast, clear selection in less than 50 years of breeding was observed for alleles in biosynthetic pathways important for market class-specific traits such as pigmentation and carbohydrate composition. Although improvement and diversification for distinct market classes was observed through whole-genome analysis of historic and current potato lines, an increased rate of gain from selection will be required to meet growing global food demands and challenges due to climate change. Understanding the genetic basis of diversification and trait improvement will allow for more rapid genome-guided improvement of potato in future breeding efforts.

Evaluation of Potato Tuber Moth (Lepidoptera: Gelechiidae) Resistance in Tubers of Bt-cry5 Transgenic Potato Lines

Abstract The potato tuber moth, Phthorimaea operculella (Zeller), in tropical and subtropical countries, is the most destructive pest of potato, Solanum tuberosum L. The larvae attack foliage and tubers in the field and in storage. The purpose of this study was to evaluate the efficacy of a Bt-cry5 transgene to control the potato tuber moth in tuber tissues. Tuber bioassays using stored (11-12 mo old) and newly harvested tubers of Bt-cry5-Lemhi Russet and Bt-cry5-Atlantic potato lines showed up to 100% mortality of 1st instars. Mortality was lowest in the newly harvested tubers of Bt-cry5-Atlantic lines (47.1–67.6%). Potato tuber moth mortality was 100% in the Bt-cry5-Spunta lines that were transformed with Bt-cry5 gene controlled by the CaMV 35S promoter (pBIML5 vector) and in 2 of 3 lines transformed with Bt-cry5 gene controlled by the Gelvin super promoter (pBIML1vector). The transgenic Spunta lines expressing Bt-cry5 controlled by the patatin promoter (pBMIL2 vector) showed the lowest tuber moth mortality (25.6 and 31.1%). The Bt-cry5 transgenic lines with high tuber expression of B. thuringiensis have value in an integrated pest management system to control potato tuber moth.

Evaluation of wildSolanum species for resistance to the US-8 genotype ofPhytophthora infestans utilizing a fine-screening technique

Greenhouse assays were carried out to characterizeSolanum accessions previously reported to be late blight resistant and to identify individuals within the accessions with high levels of resistance to late blight. Foliage of wildSolanum species accessions or hybrid cultivated potato x wild species (diploid or tetraploid) from Mexico, Russia, or South America was inoculated withPhytophthora infestans (US-8) and was scored for severity of foliar symptoms at 7 and 14 days after inoculation (DAI). Mexican accessions (S. brachycarpum,S. pinnatisectum,S. guerreroense, andS.fendleri) were significantly more resistant (1.1% infection at 14 DAI) than either Russian (59.1% infection at 14 DAI) or South American (53.4% infection at 14 DAI) accessions. Moreover, the genotypes within the Mexican accessions tended to be more uniformly resistant whereas genotypes within the Russian and South American accessions tended to segregate for resistance. The more resistant genotypes of Russian and South American accessions (176/546) were retested, and 56 genotypes were identified as having potential for use in potato breeding programs for resistance to late blight. Fifty percent of the selected genotypes were fromS. microdontum accessions PI498124, PI595510, and PI595511. The potential of these species for incorporation into breeding programs for late blight resistance is discussed.ResumenUn experimente fue conducido en invernadero para la seleción de genoti.pos entre accesiones deSolanum previamente identificados como resistentes al tizón tardÌo de la papa (Phytophthora infestans). El foliaje de accesiones de especies silvestres o hibridos entre cultivares de papa y especies silvestres (diploide y tetraploide) de MÈxico, Rusia y America del Sur fue inoculado con el genotipo US-8 de P.infestans. El porcentaje del foliage infectado fue evaluado a los 7 y 14 dias despues de la inoculación. Las accesiones de MÈxico (S. brachycarpum,S. pinnatisectum,S. guerreroense yS. fendleri) fueron significativamente mas resistentes al tizón tardÌo a los 14 dias depues de la inoculación (1,1%) que las accesiones de Rusia (59,1% ) o de America der Sur (53,4% ). Tambien, la resistencia al tizón tardÌo fue uniformemente distribuida al nivel de genotipo en las accesiones de MÈxico, pero segregación para resistancia ocurrio en accesiones de Rusia y de America del Sur. Selecciones hechas en las accesiones de Rusia y de America del Sur (176/546) fueron reevaluadas en 1998 y resultarón en 56 genotipos que pueden ser utilizados en un programa de mejoramiento para resistencia ai tizón tardÌo de la papa. Un total de 50% de los genotipos seleccionados sonSolanum microdontum accesiones PI 498124, PI 595510 y PI 595511. Tambien se discuten las posibilidades del uso y las estrategias de mejoramiento para la utilizatión de estas especies en un programa de resistencia al tizón tardÌo de la papa.

An applied fingerprinting system for cultivated potato using simple sequence repeats

The ability to quickly and accurately identify potato (Solanum tuberosum L.) clones is important to potato-breeding programs, seed and commercial potato growers, and marketing and utilization of potato cultivars. Since 1990, the Michigan State University Potato Breeding and Genetics Program has used an isozyme-based fingerprinting system to identify potato cultivars. Isozyme analysis is an economical and effective means of discriminating potato clones; however, isozyme analysis requires fresh, healthy tuber or leaf tissue. DNA-based fingerprinting using simple sequence repeats (SSRs or microsatellites) has been shown to discriminate between potato clones. The objective of this study was to identify the most useful SSR primer pairs that accurately and efficiently distinguish clones for an applied fingerprinting system of cultivated potato. SSR primer pairs with high polymorphism were selected from previous tetraploid potato studies. DNA isolated from 17 potato clones representing round-white, russet, and red market classes were visualized on both polyacrylamide (PAGE) and agarose gel systems. Polymorphism was observed in all 18 primer combinations on PAGE and 14 using agarose gel electrophoresis. All 17 cultivars were discriminated on PAGE with various combinations of two primer pairs: STIIKA using STACCAS3, STIN-HWI, or STM0031; and STACCAS3 using STGBSS1, POTM1-2, STM1104, or STM0031. The combination of STM0019, STM0031, STGBSS1, and POTM1-2 was able to differentiate all 17 clones using agarose gel electrophoresis. PAGE was determined to be the preferred system for variety identification, but agarose gel electrophoresis can be used to differentiate lines when specific varietal comparisons are needed. In addition, five different DNA source tissue types were evaluated (fresh foliar, freeze-dried foliar, fresh tuber, freeze-dried tuber epidermis, and freeze-dried tuber tissue). Amplification products were similar for all five tissue sources used for DNA isolation. This ability to isolate DNA from freeze-dried tissue will allow cultivar identification when fresh tissue is not available. The SSR primer pairs presented here can be used as a practical fingerprinting system for cultivated potato identification.ResumenLa posibilidad de identificatión rápida y exacta de clones de papa (Solanum tuberosum L.), es importante para los programas de mejoramiento, para los productores de semilla y papa de consumo, lo mismo que para la comercialización y utilizatión de los cultivares de papa. Desde 1990, el Programa de Mejoramiento y Genética de la Universidad del Estado de Michigan ha utilizado un sistema de tipificación genética para identificar cultivares de papa basado en isoenzimas. El análisis de isoenzimas es un método económico y efectivo para diferenciar clones de papa. Sin embargo, este análisis requiere de tejido fresco y sano de hoja o de tubérculo. Con la tipificación genética basada en el uso de ADN y utilizando repeticiones de secuencias simples (SSRs o microsatélites) se ha demostrado la existencia de diferencias entre clones de papa. El objetivo del presente estudio fue identificar los pares de iniciadores SSR más útiles para distinguir clones de papa cultivada en forma precisa y eficiente por medio de un sistema de tipificación genética aplicada. A partir de estudios previos de papa tetraploide se seleccionaron pares de iniciadores SSR con polimorfismo amplio. El ADN aislado de 17 clones de papa representantes de las clases comerciales redondo bianco, rosado y rojo se hizo evidente tanto en el sistema de poliacrilamida (PAGE) como en el de gel de agarosa. Con el sistema PAGE se observó polimorfismo en todas las combinaciones de los 18 iniciadores, y en 14 cuando se utilizó electroforesis en gel de agarosa. Los 17 cultivares fueron diferenciados en PAGE con varias combinaciones de dos pares de iniciadores: STIKA utilizando STACCAS3, STINHWI o STM0031 y STACCAS utilizando STGBSS1, POTM1-2, STM1104, o STM0031. La combinación de STM0019, STM0031, STGBSS1 y POTM1-2 fue capaz de diferenciar los 17 clones utilizando electroforesis en gel de agarosa. Se estableció que PAGE es el sistema preferido para la identification de variedades, pero se puede utilizar la electroforesis en gel de agarosa para diferenciar lineas, cuando se necesitan comparaciones varietales especificas. Además se evaluaron cinco fuentes de tejido (foliar fresco, foliar seco congelado, tubérculo fresco, epidermis de tubérculo seco congelado, tubérculo seco congelado). Los productos de amplificacion fueron similares para las cinco fuentes de tejido para el aislamiento de ADN. Esta facultad para aislar ADN de tejido seco congelado permitirá la identificacion de cultivares, cuando no se disponga de tejido fresco. Los pares del iniciador SSR presentados aqui se pueden utilizar como un sistema práctico de tipificación genética para la identificación de la papa cultivada.

Genetic linkage mapping of economically important traits in cultivated tetraploid potato (Solanum tuberosum L.)

The objective of this study was to construct a single nucleotide polymorphism (SNP)-based genetic map at the cultivated tetraploid level to locate quantitative trait loci (QTL) contributing to economically important traits in potato (Solanum tuberosum L.). The 156 F1 progeny and parents of a cross (MSL603) between “Jacqueline Lee” and “MSG227-2” were genotyped using the Infinium 8303 Potato Array. Furthermore, the progeny and parents were evaluated for foliar late blight reaction to isolates of the US-8 genotype of Phytophthora infestans (Mont.) de Bary and vine maturity. Linkage analyses and QTL mapping were performed using a novel approach that incorporates allele dosage information. The resulting genetic maps contained 1972 SNP markers with an average density of 1.36 marker per cM. QTL mapping identified the major source of late blight resistance in “Jacqueline Lee.” The best SNP marker mapped ∼0.54 Mb from a resistance hotspot on the long arm of chromosome 9. For vine maturity, the major-effect QTL was located on chromosome 5 with allelic effects from both parents. A candidate SNP marker for this trait mapped ∼0.25 Mb from the StCDF1 gene, which is a candidate gene for the maturity trait. The identification of markers for P. infestans resistance will enable the introgression of multiple sources of resistance through marker-assisted selection. Moreover, the discovery of a QTL for late blight resistance not linked to the QTL for vine maturity provides the opportunity to use marker-assisted selection for resistance independent of the selection for vine maturity classifications.

Jacqueline Lee: A late-blight-resistant tablestock variety

Jacqueline Lee has a bright golden skin, yellow flesh, attractive oval shape, and excellent cooking qualities that make it suitable for tablestock use. In addition, it has been determined to have a high level of foliar resistance to the US-8 genotype ofPhytophthora infestans (Mont.) de Bary under Michigan field and greenhouse conditions. It is a full-season variety with vine maturity similar to Snowden. Jacqueline Lee has high yield potential. It has a high set (14–16 tubers/hill) of medium sized tubers that leads to 65%–80% marketable yield. Tubers are low in internal and external defects. It has a scab reaction similar to Atlantic, and the tubers have been observed to have a long dormancy.ResumenJacqueline Lee es una variedad de papa de piel amarilla brillante para consumo fresco, tubérculos de forma oval muy atractivos y de excelente calidad de cocimiento. Esta variedad tiene altos niveles de resistencia foliar al genotypo US-8 de Phytophthora infestans (Mont.) de Bary, tanto en invernadero como en condiciones de campo, esta evaluacion fue hecha en Michigan, USA. Jacqueline Lee es una variedad con alto potencial de rendimiento y maduración media tardia similar a Snowden. Produce un gran número de tubérculos (14-16 tubérculos/semilla) de tamaño mediano, de los cuales 65-80% son tubérculos comercializables. Los tubérculos tienen resistencia a defectos internos y externos. Tambien tienen resistencia a la sarna (Streptomyces scabies Thaxter) similar a la variedad Atlantic y dormancia larga.

Greenhouse and Field Nursery Evaluation for Potato Common Scab Tolerance in a Tetraploid Population

Potato common scab (Streptomyces scabies (Thaxt.) Waksman & Henrici) is a major disease of potato (Solanum tuberosum L.), due to the unmarketability of affected tubers. For identification of the most common scab-tolerant material, and for developing molecular markers for common scab tolerance, more information is needed on the genetic basis of common scab tolerance. Phenotyping common scab susceptibility is difficult because of the large variability in disease symptoms among tubers from a single plant, ranging from no common scab to severe pits. Two years of field data were collected for scab reaction on a segregating tetraploid population (MSL603, 160 individuals). Continuous variation in common scab susceptibility phenotype was observed among the progeny, with a normal distribution suggesting common scab disease phenotype is a genetically complex trait. Transgressive segregants were also observed, but they are skewed toward susceptibility. A greenhouse-based screening procedure was evaluated to discern tolerant from susceptible potato lines. A subset of ten individuals from this population were selected (five resistant, five susceptible). For the greenhouse study, soil was inoculated with a pathogenic S. scabies strain MSDPZ at a concentration of 3 X 108 CFU/ml. This greenhouse assay effectively discerned tolerant and susceptible individuals. There was a moderate correlation between the greenhouse study and the field trial. The greenhouse assay may provide information that would complement field data in identifying resistant clones.ResumenLa sarna común (Streptomyces scabies (Thaxt.) Waksman & Henrici) en una enfermedad muy importante de la papa (Solanum tuberosum L.) debido a que los tubérculos afectados no son comerciables. Para la identificación del material más tolerante a la sarna común y para desarrollar marcadores moleculares para tolerancia a la enfermedad se necesita más información sobre la base genética de la tolerancia. La fenotipificación de la susceptibilidad a la sarna común es difícil por la gran variabilidad en los síntomas de la enfermedad entre los tubérculos de una misma planta, que varían desde la ausencia de sarna a presencia de hoyos profundos en los tubérculos. Se colectaron datos durante dos años sobre la reacción a la sarna en una población tetraploide (MSL603, 160 individuos). La variación continua del fenotipo susceptible a la sarna común fue observada entre la progenie, con una distribución normal que sugiere que el fenotipo de la enfermedad es una característica genéticamente compleja. También se observó variación transgresiva, pero sesgada hacia la susceptibilidad. Un procedimiento de tamizado en invernadero fue evaluado para diferenciar las líneas de papa tolerantes de las susceptibles. Un subgrupo de 10 individuos de la población fue seleccionado (cinco resistentes, cinco susceptibles). Para el estudio de invernadero, el suelo fue inoculado con una cepa patogénica MSDPZ de S. scabies a una concentración de 3 x 108 UFC/ml. Este ensayo de invernadero distinguió efectivamente individuos tolerantes y susceptibles. Hubo una moderada correlación entre los estudios de invernadero y la prueba de campo. La prueba de invernadero puede proveer información que complementaría los datos de campo para identificar clones resistentes.

Missaukee: A Round White Potato Variety Combining Chip-Processing With Resistance to Late Blight, Verticillium Wilt and Golden Cyst Nematode

Previously identified as breeding line MSJ461-1, Missaukee is a round white chip processing potato variety resulting from a cross between Tollocan and NY88 and has foliar resistance to potato late blight (Phytophthora infestans de Bary). This variety has an attractive round shape and mildly netted, bright skin. Seven years of field testing in Michigan indicate that the yield of total marketable tubers in Missaukee is similar to that of Snowden. However, Missaukee has a lower incidence of internal defects than Snowden. Specific gravity ranged from 1.069 to 1.086 in Michigan trials and out-of-the-field chip scores were similar to those of Snowden. Missaukee showed some resistance to Verticillium wilt in 2-years of trials. DNA marker and greenhouse tests indicate that Missaukee is also resistant to the golden cyst nematode (Globodera rostochiensis Woll) pathotype Ro1.ResumenPreviamente identificada como la línea de mejoramiento MSJ461-1, Missaukee es una variedad de papa redonda, blanca, para proceso de fritura, como resultado de la cruza entre Tollocan y NY88 y tiene resistencia foliar al tizón tardío de la papa (Phytophthora infestans de Bary). Esta variedad tiene una forma redonda atractiva y piel brillante, ligeramente con red. Siete años de pruebas de campo en Michigan indican que el rendimiento total de tubérculos comerciales en Missaukee es similar al de Snowden. No obstante, Missaukee tiene mas baja incidencia de defectos internos que Snowden. La gravedad específica fluctuó entre 1.069 a 1.086 en los ensayos de Michigan y fuera del campo los valores de fritura fueron similares a los de Snowden. Missaukee mostró alguna resistencia al marchitamiento por Verticillium en ensayos de dos años. Los marcadores de DNA y las pruebas de invernadero indican que Missaukee también es resistente al nematodo dorado de quiste (Globodera rostochiensis Woll) patotipo Ro1.

The Contribution of the Solanaceae Coordinated Agricultural Project to Potato Breeding

Potato variety development in the USA involves extensive interaction between public sector scientists and state and national grower organizations to test and commercialize new varieties. Historically, breeding and germplasm development efforts have relied upon phenotypic evaluations to select and advance germplasm. The goal of the US Department of Agriculture (USDA)-funded Solanaceae Coordinated Agricultural Project (SolCAP) was to translate genomic resources into tools that can be used by breeders and geneticists. A major outcome of this project was the development of a genome-wide single nucleotide polymorphism (SNP) array that can be used to evaluate elite potato breeding germplasm. This array was utilized to genotype numerous biparental tetraploid and diploid populations, a diversity panel, and a core collection of Solanum species. The SNP array provides a marker density sufficient to generate genetic maps to identify numerous quantitative trait loci (QTLs) for agronomic, quality, and disease resistance traits. Many new diploid and tetraploid genetic maps have been constructed and used to identify numerous QTLs for important traits. We used SNP markers to assess relationships among germplasm, fingerprint varieties, and identify candidate genes. The Infinium 8303 Potato Array provides a common set of SNP markers that can reliably be used for mapping, germplasm assessment, and fingerprinting. This array has also been a useful tool to advance our understanding of the potato genome. Furthermore, breeders are mapping QTLs across numerous populations that will expand our understanding of economically important traits and lead to marker-assisted selection and breeding, and ultimately improved varieties.