A de novo genome assembly of Solanum verrucosum Schlechtendal, a Mexican diploid species geographically isolated from other diploid A-genome species of potato relatives.

There are over 100 known species of cultivated potatoes and their wild relatives. Many of these species, including cultivated potatoes, share the A genome; these species are mainly distributed in South America and are reproductively isolated from Mexican diploid species. The only diploid A-genome species distributed in Mexico is Solanum verrucosum Schlechtendal, which is also a maternal progenitor of Mexican polyploid species. In this study, we constructed a high-quality de novo assembly of the S. verrucosum genome using PacBio long-read sequencing and Hi-C scaffolding technologies. A monohaploid clone (2n = x = 12) of S. verrucosum was used to reduce assembly difficulty due to the heterozygous nature of the species. The final sequence assembly consisted of 780.2 Mb of sequence, 684.0 Mb of which were anchored to the 12 chromosomes, with a scaffold N50 of 55.2 Mb. Putative centromeres were identified using publicly available data obtained via chromatin immunoprecipitation sequencing against a centromere-specific histone 3 protein. Transposable elements accounted for approximately 61.8% (482.1 Mb) of the genome, and 46,904 genes were functionally annotated. High gene synteny and similarity were revealed among the genomes of S. verrucosum, Solanum commersonii, Solanum chacoense, Solanum phureja, Solanum tuberosum, and Solanum lycopersicum. The reference-quality S. verrucosum genome will provide new insights into the evolution of Mexican polyploid species and contribute to potato breeding programs.

Discovery of a novel mitochondrial DNA molecule associated with tetrad pollen sterility in potato.

BACKGROUND: Tetrad sterility in potato is caused by a specific cytoplasm, called TSCsto, derived from the Mexican wild tetraploid species Solanum stoloniferum. Different S. stoloniferum accessions crossed as females with S. tuberosum resulted in 12 fertile hybrids and 27 sterile hybrids exhibiting tetrad sterility. RESULTS: Whole-mitochondrial-genome sequencing was performed for two fertile hybrids and three hybrids exhibiting tetrad sterility. Two to seven contigs, with the total assembly lengths ranging from 462,716 to 535,375 bp, were assembled for each hybrid. Unlike for the reference mitochondrial genome (cv. Désirée), two different recombinant-type contigs (RC-I and RC-II) were identified. RC-I featured by the rpl5-ψrps14 gene joined to the nad6 gene, generating a novel intergenic region. Using a PCR marker (P-3), we found that this intergenic region occurred exclusively in interspecific hybrids exhibiting tetrad sterility and in their parental S. stoloniferum accessions. A part of this intergenic sequence was expressed in the pollen. From a large survey in which P-3 was applied to 129 accessions of 27 mostly Mexican wild species, RC-I was found in diploid S. verrucosum and polyploid species. From eight accessions of S. verrucosum used as females, 92 interspecific hybrids were generated, in which only those carrying RC-I exhibited tetrad sterility. CONCLUSIONS: RC-I was clearly associated with tetrad sterility, and the RC-I-specific intergenic region likely contains a causal factor of tetrad sterility.

Cytoplasmic genome types of European potatoes and their effects on complex agronomic traits.

BACKGROUND: Various wild species germplasm has been used in European potato breeding since the first introduction of potato (Solanum tuberosum L.) to Europe. As the plant cytoplasmic genome including chloroplast and mitochondrial genomes is transmitted only through the maternal parent, cytoplasmic markers are useful tools in breeding programs to determine cytoplasmic genome types and to trace maternal ancestors. The potato cytoplasmic genome can be distinguished into six distinct types (M, P, A, W, T, and D). Male sterility was found in genotypes with S. demissum-derived D-type cytoplasm and S. stoloniferum-derived W/γ-type cytoplasm. These wild species were frequently used to incorporate superior pathogen resistance genes. As a result, the percentage of these two types is increasing unintentionally in the European germplasm pool. Other than cytoplasmic male sterility, little is known about effects of the cytoplasmic genome on complex agronomic traits in potato. RESULT: The cytoplasm types of 1,217 European potato cultivars and breeding clones were determined with type specific DNA markers. Most frequent were T- (59.4 %), D- (27.4 %), and W- (12.2 %) type cytoplasm, while A- (0.7 %) and M-type cytoplasm (0.3 %) was rare and P-type cytoplasm was absent. When comparing varieties with breeding clones, the former showed a relatively higher frequency of T-type and lower frequency of D- and W-type cytoplasm. Correlation analysis of cytoplasm types and agronomic data showed that W/γ-type cytoplasm was correlated with increased tuber starch content and later plant maturity. Correlation with quantitative resistance to late blight was observed for D-type and M-type cytoplasm. Both cytoplasm types had a positive effect on resistance. CONCLUSION: This study revealed and quantified the cytoplasmic diversity in the European potato germplasm pool. Knowledge of cytoplasm type is important for maintaining genetic diversity and managing the male sterility problem in breeding programs. This is the first comprehensive study to show correlations of distinct cytoplasmic genomes with complex agronomic traits in potato. Correlations particularly with tuber starch content and resistance to late blight provided new knowledge on cytoplasmic effects on these important traits, which can be exploited for genetic improvement of potato.

Genomic architecture of potato resistance to Synchytrium endobioticum disentangled using SSR markers and the 8.3k SolCAP SNP genotyping array.

BACKGROUND: The soil borne, obligate biotrophic fungus Synchytrium endobioticum causes tumor-like tissue proliferation (wart) in potato tubers and thereby considerable crop damage. Chemical control is not effective and unfriendly to the environment. S. endobioticum is therefore a quarantined pathogen. The emergence of new pathotypes of the fungus aggravate this agricultural problem. The best control of wart disease is the cultivation of resistant varieties. Phenotypic screening for resistant cultivars is however time, labor and material intensive. Breeding for resistance would therefore greatly benefit from diagnostic DNA markers that can be applied early in the breeding cycle. The prerequisite for the development of diagnostic DNA markers is the genetic dissection of the factors that control resistance to S. endobioticum in various genetic backgrounds of potato. RESULTS: Progeny of a cross between a wart resistant and a susceptible tetraploid breeding clone was evaluated for resistance to S. endobioticum pathotypes 1, 2, 6 and 18 most relevant in Europe. The same progeny was genotyped with 195 microsatellite and 8303 single nucleotide polymorphism (SNP) markers. Linkage analysis identified the multi-allelic locus Sen1/RSe-XIa on potato chromosome XI as major factor for resistance to all four S. endobioticum pathotypes. Six additional, independent modifier loci had smaller effects on wart resistance. Combinations of markers linked to Sen1/RSe-XIa resistance alleles with one to two additional markers were sufficient for obtaining high levels of resistance to S. endobioticum pathotypes 1, 2, 6 and 18 in the analyzed genetic background. CONCLUSIONS: Potato resistance to S. endobioticum is oligogenic with one major and several minor resistance loci. It is composed of multiple alleles for resistance and susceptibility that originate from multiple sources. The genetics of resistance to S. endobioticum varies therefore between different genetic backgrounds. The DNA markers described in this paper are the starting point for pedigree based selection of cultivars with high levels of resistance to S. endobioticum pathotypes 1, 2, 6 and 18.

Pollen transcriptome analysis of Solanum tuberosum (2n = 4x = 48), S. demissum (2n = 6x = 72), and their reciprocal F1 hybrids.

KEY MESSAGE: Pollen mRNAs were different in reciprocal F 1 hybrids, which were probably caused by a cytoplasm-nuclear chromosomal genes interaction. We have found reciprocal differences in crossability between F1 hybrids of Solanum tuberosum (T) and a Mexican wild potato species S. demissum (D). When the reciprocal hybrids were crossed as pollen parents with S. demissum, a significantly higher berry-setting rate was obtained in TD compared with DT. In this study, we performed a whole-genome transcript analysis of the pollen mRNA using a high-throughput sequencer. We obtained 12.6 billion bases that were aligned into 13,020 transcripts with 9,366 loci. All possible genetic modes were observed between the parents and their progeny, where over-dominance and under-recessive types were relatively frequent (15.7 and 15.3 %, respectively). We found that 59.1 % of transcripts were more abundant in TD and over fourfold higher transcription levels were found in 66 TD transcripts and three DT transcripts. A higher proportion of over-dominance and a lower proportion of under-recessive transcription types were also observed in TD. The percentage contributions of multiple transcripts at the same locus varied greatly and were transcribed differently between species. In the new allelic combinations created by hybridization, approximately three-fourth of the transcripts had intermediate percentage contributions between the parents but no differential transcription patterns were apparent between the reciprocal hybrids. A broad spectrum of functionally different nuclear genes was over-represented in TD pollen, some of which were directly related to pollen behavior. Since TD and DT pollen had the same composition of nuclear genes, a cytoplasm-nuclear chromosomal genes interaction is suggested for the cause of transcriptional and phenotypic differences between reciprocal hybrids.

Development of a rapid identification method for potato cytoplasm and its use for evaluating Japanese collections.

The cytoplasm of potatoes, characterized by the presence of T-type chloroplast DNA and ß-type mitochondrial DNA, is sensitive to nuclear chromosomal genes that contribute to various types of male sterility. Past breeding efforts with various potato varieties have resulted in several different cytoplasms other than T/ß. Varieties with Solanum stoloniferum-derived cytoplasm (W/γ) show complete male sterility, while those with S. demissum-derived cytoplasm (W/α) produce abundant, but non-functional pollen. Thus, identification of cytoplasmic types is important for designing efficient mating combinations. To date, only T-type chloroplast DNA can be accurately identified by a PCR marker. Here, we report a rapid identification technique by multiplex PCR, followed by restriction digestion with BamHI in one reaction tube, and propose a new nomenclature for potato cytoplasm types (T, D, P, A, M, and W). Using this new technique, our collections of 748 genotypes, including 84 Japanese named varieties, 378 breeding lines and 26 landraces, and 260 foreign varieties and breeding lines, were grouped into cytoplasm types: T (73.9 %), D (17.4 %), P (4.5 %), A (1.5 %), M (0.3 %), and W (2.4 %). The utility of this marker system for breeding is discussed.

Comparative differentiation in mitochondrial and chloroplast DNA among cultivated potatoes and closely related wild species.

A total of 476 accessions of seven cultivated and 32 wild potato species previously characterized by nuclear DNA (nDNA) and chloroplast DNA (ctDNA) marker analyses were employed to the mitochondrial DNA (mtDNA) marker analysis. Fourteen simple sequence repeat (SSR) markers with mononucleotide repeat regions were developed from the potato mtDNA, although their variability was extremely low. Six mtDNA markers including three developed SSR markers disclosed 40 banding patterns that discriminated 63 different mtDNAs. For the same set of samples, 72 ctDNA banding patterns discriminated 129 different ctDNAs. Consequently, 164 haplotypes were distinguished. The correlation between ctDNA and mtDNA differentiation was positive (r = 0.226), but poor when compared with that between ctDNA and nDNA (r = 0.415), which likely lowered the utility of mtDNA polymorphisms in evaluating relationships among these species. Nevertheless, a finding of a unique mtDNA type in all T-type ctDNA holders (S. tuberosum and S. tarijense) strongly supports S. tarijense functioned as a maternal ancestor of potato.