David De Koeyer

Physiological and molecular adaptations to drought in Andean potato genotypes

The drought stress tolerance of two Solanum tuberosum subsp. andigena landraces, one hybrid (adg×tbr) and Atlantic (S. tuberosum subsp. tuberosum) has been evaluated. Photosynthesis in the Andigena landraces during prolonged drought was maintained significantly longer than in the Tuberosum (Atlantic) line. Among the Andigena landraces, ‘Sullu’ (SUL) was more drought resistant than ‘Negra Ojosa’ (NOJ). Microarray analysis and metabolite data from leaf samples taken at the point of maximum stress suggested higher mitochondrial metabolic activity in SUL than in NOJ. A greater induction of chloroplast-localized antioxidant and chaperone genes in SUL compared with NOJ was evident. ABA-responsive TFs were more induced in NOJ compared with SUL, including WRKY1, mediating a response in SA signalling that may give rise to increased ROS. NOJ may be experiencing higher ROS levels than SUL. Metabolite profiles of NOJ were characterized by compounds indicative of stress, for example, proline, trehalose, and GABA, which accumulated to a higher degree than in SUL. The differences between the Andigena lines were not explained by protective roles of compatible solutes; hexoses and complex sugars were similar in both landraces. Instead, lower levels of ROS accumulation, greater mitochondrial activity and active chloroplast defences contributed to a lower stress load in SUL than in NOJ during drought.

Genes driving potato tuber initiation and growth: identification based on transcriptional changes using the POCI array

The increasing amount of available expressed gene sequence data makes whole-transcriptome analysis of certain crop species possible. Potato currently has the second largest number of publicly available expressed sequence tag (EST) sequences among the Solanaceae. Most of these ESTs, plus other proprietary sequences, were combined and used to generate a unigene assembly. The set of 246,182 sequences produced 46,345 unigenes, which were used to design a 44K 60-mer oligo array (Potato Oligo Chip Initiative: POCI). In this study, we attempt to identify genes controlling and driving the process of tuber initiation and growth by implementing large-scale transcriptional changes using the newly developed POCI array. Major gene expression profiles could be identified exhibiting differential expression at key developmental stages. These profiles were associated with functional roles in cell division and growth. A subset of genes involved in the regulation of the cell cycle, based on their Gene Ontology classification, exhibit a clear transient upregulation at tuber onset indicating increased cell division during these stages. The POCI array allows the study of potato gene expression on a much broader level than previously possible and will greatly enhance analysis of transcriptional control mechanisms in a wide range of potato research areas. POCI sequence and annotation data are publicly available through the POCI database (http://pgrc.ipk-gatersleben.de/poci).

Application of high-resolution DNA melting for genotyping and variant scanning of diploid and autotetraploid potato

The ideal marker system for tetraploid potato would be dosage-sensitive and have the ability to distinguish heterozygous genotypes with multiple haplotypes within the genomic region targeted by the marker. The objective of this study was to evaluate the utility of high-resolution DNA melting (HRM) for genotyping and polymorphism detection in diploid and tetraploid potato. Amplicon scanning, unlabelled probe, and short amplicon assays were developed for four candidate genes affecting tuber skin and flesh colour, and starch, and a marker linked to nematode resistance. Genotyping a set of 95 potato clones revealed several examples of clones with three distinct haplotypes. Combined probe and amplicon analysis identified between 29 and 44 unique genotypes for the same assays. Assays developed for four of the five target genes are suitable for marker-assisted selection in potato breeding programs. This study illustrates the use of HRM in potato genetics. Further advances in the technology and associated data analysis should make HRM a useful tool for basic and applied studies of potato.

Finding the perfect potato: using functional genomics to improve disease resistance and tuber quality traits

One of the best known plant disease outbreaks of all time was the Irish potato famine of 1845 to 1847 when the late blight pathogen, Phytophthora infestens, devastated the potato crop and caused more than one million deaths in Ireland. Potatoes are still plagued by disease but luckily most countries do not depend on potato as much as they did in Ireland in the 1800s. Producing disease-free potatoes, however, has come at a huge cost. To combat major diseases, such as late blight and common scab, 64 × 106 kg of pesticides are sprayed on potato fields each year. The costs associated with these measures are not only financial; environmental costs, although less easy to quantify, are significant and include negative impacts on natural ecosystems and the contamination of groundwater, lakes, and rivers. Our search for the “perfect” potato, i.e., one that has good processing qualities and disease resistance, has gained new momentum with the emergence of genomic technologies. Through functional genomics we will gain a better understanding of the genes responsible for tuber quality traits and those responsible for disease resistance. With a collection of desirable genes in mind, we can again use genomics as a diagnostic tool to search for these genes in the wide variety of potatoes around the world and to follow their transfer by classical breeding. This paper describes a research program currently underway in Canada that uses functional genomics to improve the potato.

Implications of miR166 and miR159 induction to the basal response mechanisms of an andigena potato (Solanum tuberosum subsp. andigena) to salinity stress, predicted from network models in Arabidopsis

MicroRNA (miRNA) mediated changes in gene expression by post-transcriptional modulation of major regulatory transcription factors is a potent mechanism for integrating growth and stress-related responses. Exotic plants including many traditional varieties of Andean potatoes (Solanum tuberosum subsp. andigena) are known for better adaptation to marginal environments. Stress physiological studies confirmed earlier reports on the salinity tolerance potentials of certain andigena cultivars. Guided by the hypothesis that certain miRNAs play important roles in growth modulation under suboptimal conditions, we identified and characterized salinity stress-responsive miRNA-target gene pairs in the andigena cultivar Sullu by parallel analysis of noncoding and coding RNA transcriptomes. Inverse relationships were established by the reverse co-expression between two salinity stress-regulated miRNAs (miR166, miR159) and their target transcriptional regulators HD-ZIP-Phabulosa/Phavulota and Myb101, respectively. Based on heterologous models in Arabidopsis, the miR166-HD-ZIP-Phabulosa/Phavulota network appears to be involved in modulating growth perhaps by mediating vegetative dormancy, with linkages to defense-related pathways. The miR159-Myb101 network may be important for the modulation of vegetative growth while also controlling stress-induced premature transition to reproductive phase. We postulate that the induction of miR166 and miR159 under salinity stress represents important network hubs for balancing gene expression required for basal growth adjustments.

Decreased defense gene expression in tolerance versus resistance to Verticillium dahliae in potato

Verticillium dahliae Kleb., a soil-borne fungus that colonizes vascular tissues, induces wilting, chlorosis and early senescence in potato. Difference in senescence timing found in two diploid potato clones, 07506-01 and 12120-03, was studied and genetic variation in response to V. dahliae infection was identified as a causal factor. The clone, 07506-01, was infected with V. dahliae but did not develop symptoms, indicating tolerance to the pathogen. The other diploid clone, 12120-03 had low levels of pathogen with infection and moderate symptoms indicating partial resistance. 07506-01 was found to carry two susceptible alleles of the Ve2 gene and 12120-03 carried one Ve2 resistant and one susceptible allele. Infected leaves of the two clones were compared using gene expression profiling with the Potato Oligonucleotide Chip Initiative (POCI) microrarray. The results provide further evidence for differences in response of the two clones to infection with V. dahliae. Chlorophyll biosynthesis was higher in the tolerant 07506-01 compared to partially resistant 12120-03. On the other hand, expression of fungal defense genes, Ve resistance genes and defense phytohormone biosynthetic enzyme genes was decreased in 07506-01 compared to 12120-03 suggesting defense responses were suppressed in tolerance compared to resistance. Transcription factor gene expression differences pointed to the WRKY family as potential regulators of V. dahliae responses in potato.

Comparative metabolite profiling of Solanum tuberosum against six wild Solanum species with Colorado potato beetle resistance.

The Colorado potato beetle Leptinotarsa decemlineata (Say) (CPB) is a coleopteran herbivore that feeds on the foliage on Solanum species, in particular, potato. Six resistant wild Solanum species were identified, and two of these species had low levels of glycoalkaloids. Comparative analysis of the untargeted metabolite profiles of the foliage using UPLC-qTOF-MS was done to find metabolites shared between the wild species but not with Solanum tuberosum (L.) to identify resistance-related metabolites. It was found that only S. tuberosum produced the triose glycoalkaloids solanine and chaconine. Instead, the six wild species produced glycoalkaloids that shared in common tetrose sugar side chains. Additionally, there were non-glycoalkaloid metabolites associated with resistance including hydroxycoumarin and a phenylpropanoid, which were produced in all wild species but not in S. tuberosum.

Continued response through seven cycles of recurrent selection for grain yield in oat (Avena sativa L.)

Long-term selection experiments provide germplasm to study the effects of selection in a closed population. Recurrent selection to enhance grain yield in oat has been ongoing at the University of Minnesota since 1968. The objectives of this study were: (i) estimate the GCA and SCA effects for three agronomic traits in the seventh cycle of selection, (ii) assess the effect of the current methods of selection on parental contribution and unselected traits, and (iii) determine the direct and indirect responses to seven cycles of recurrent selection for grain yield. Progeny of the Cycle 6 parents and parents for Cycles 0 through 7 were grown in two separate tests. Grain yield, heading date and plant height were evaluated in each test. Grain yield was increased by 21.7% after seven cycles of selection. Evaluation of Cycle 6 progeny showed that GCA effects were significant for all three traits studied, and SCA effects were significant only for grain yield. Four Cycle 6 parents did not have any progeny selected as Cycle 7 parents. Results from this study indicate that long term recurrent selection has continued to increase grain yield. Alternative selection strategies may be necessary to maintain the genetic variability in this population, particularly when improvement of secondary traits is required.

Differences between the Bud End and Stem End of Potatoes in Dry Matter Content, Starch Granule Size, and Carbohydrate Metabolic Gene Expression at the Growing and Sprouting Stages

Potatoes usually have the tuber bud end dominance in growth during tuber bulking and in tuber sprouting, likely using carbohydrates from the tuber stem end. We hypothesized that the tuber bud end and tuber stem end coordination in carbohydrate metabolism gene expression is different between the bulking dominance and sprouting dominance of the tuber bud end. After comparing the growing tubers at harvest from a green vine and the stage that sprouts just started to emerge after storage of tubers at room temperature, we found the following: (1) Dry matter content was higher in the tuber stem end than the tuber bud end at both stages. (2) The starch granule size was larger in the tuber bud end than in the tuber stem end. (3) The tuber bud end had higher gene expression for starch synthesis but a lower gene expression of sucrose transporters than the tuber stem end during tuber growing. (4) The tuber stem end at the sprouting stage showed more active gene expression in both starch degradation and resynthesis, suggesting more active export of carbohydrates, than the tuber bud end. The results indicate that the starch accumulation mechanism in the tuber bud end was different between field growing and post-harvest sprouting tubers and that tubers already increased dry matter and average starch granule sizes in the tuber bud end prior to the rapid growth of sprouts.

Colorado Potato Beetle Resistance in Solanum oplocense X Solanum tuberosum Intercross Hybrids and Metabolite Markers for Selection

S. oplocense Hawkes, a wild relative of the potato S. tuberosum L. and source of resistance against the Colorado potato beetle Leptinotarsa decemlineata (Say) (CPB), was intercrossed with S. tuberosum. Backcross clones carried varying levels of resistance. Differences in foliar metabolites between resistant and susceptible clones were analyzed using liquid chromatography-mass spectrometry (LC-MS). Supervised machine learning classification methods uncorrelated shrunken centroids (USC), k-nearest neighbor (KNN) and support vector machines (SVM) were applied to develop algorithms that can classify resistant and susceptible plants using the metabolite data. Five metabolites were found to have a low error rate of prediction of CPB resistance. The five metabolites included two glycoalkaloids previously associated with resistance and susceptibility to CPB, dehydrocommersonine and solanine, respectively. Resistance was associated with a change in composition of glycoalkaloids to higher ratios of dehydrocommersonine over solanine.ResumenS. oplocense Hawkes, un pariente silvestre de la papa S. tuberosum L., y fuente de resistencia contra el escarabajo de Colorado Leptinotarsa decemlineata (Say) (CPB), se intercruzó con S. tuberosum. Los clones de la retrocruza conservaron diversos niveles de resistencia. Se analizaron las diferencias en los metabolitos foliares entre los clones resistentes y susceptibles usando espectrometría de cromatografía líquida de masas (LC-MS). Métodos supervisados de clasificación de aprendizaje de máquina no correlacionados con centroides encogidos (USC), k-cercanía de vecinos (KNN) y máquinas de respaldo de vector (SVM) se aplicaron para desarrollar algoritmos que pueden clasificar plantas resistentes y susceptibles usando los datos de los metabolitos. Se encontró que cinco metabolitos tenían un nivel bajo de error de predicción de la resistencia al CPB. Los cinco metabolitos incluyeron dos glicoalcaloides asociados previamente con resistencia y susceptibilidad al CPB, la deshidrocommersonina y la solanina, respectivamente. La resistencia se asoció con un cambio en la composición de los glicoalcaloides a altas proporciones de deshidrocommersonina sobre la solanina.