Generation of genome wide linkage maps for a wild potato and RNA-seq analysis of transgene mediated potato defense mechanisms against late blight in the tubers and foliage

Abstract

Wild potato Solanum bulbocastanum is a rich source of genetic resistance against a variety of pathogens. This project developed molecular tools and expanded biological knowledge useful for the improvement of cultivated potato (S. tuberosum) using genetic resistance from S. bulbocastanum. First, the genome structure of S. bulbocastanum relative to those of its relatives, cultivated potato and tomato (S. lycopersicon) was determined. Second, to facilitate efforts to improve cultivated potato through the introgression and deployment disease resistance derived from S. bulbocastanum, the phenotypic function and molecular mechanisms of one S. bulbocastanum disease resistance gene were explored in cultivated potato foliage and tubers. For determination of genome structure for S. bulbocastanum, Diversity Arrays Technology (DArT) was employed to generate genome wide linkage maps for the species. Employing a pseudo-testcross mapping strategy, 631 DArT markers were integrated into a composite map comprising 12 linkage groups. Our results represent an over ten-fold increase of total marker density compared to previously available genetic maps for the species. Sequencing and alignment of corresponding DArT clones to reference physical maps from tomato and cultivated potato allowed a direct comparison of marker orders between species. Overall, the S. bulbocastanum genetic maps show higher collinearity with reference potato maps than tomato maps, with seven genome regions supporting a closer phylogenetic relationship between potato and S. bulbocastanum than between tomato and S. bulbocastanum. One dominant US cultivar ‘Russet Burbank’(WT; late blight susceptible in foliage and tuber) and its RB (a late blight resistance gene derived from S. bulbocastanum) transgenic line SP2211 (+RB; late blight resistant in foliage and tuber) were compared in both tubers and foliage in their responses to late blight pathogen attack using an RNA-seq approach. In the tubers, a total of 483 million paired end Illumina RNA-seq reads were generated, representing the transcription of 29,319 potato genes. Differentially expressed genes, gene groups and ontology bins that exhibited differences between the WT and +RB lines were identified. P. infestans transcripts, including those of known effectors, were also identified. Faster and stronger activation of defense related genes, gene groups and ontology bins correlated with successful tuber resistance against P. infestans. Our results suggest that the hypersensitive response is likely a general form of resistance against the hemibiotrophic P. infestans—even in potato tubers, organs that develop below ground. In the foliage, a total of 515 million paired end RNA-seq reads were generated, representing the transcription of 29,970 genes. We compared the differences and similarities of responses to P. infestans in potato foliage and tubers. Differentially expressed genes, gene groups and ontology bins were identified to show similarities and differences in foliage and tuber defense mechanisms. Our results suggest that disease resistance gene dosage and shared biochemical pathways contribute to RB-mediated incompatible potato-P. infestans interactions in both the foliage and tubers.