Chromosome-scale reference genome assembly of a diploid potato clone derived from an elite variety

Potato (Solanum tuberosum L.) is one of the most important crops with a world-wide production of 370 million metric tons. The objectives of this study were (i) to create a high quality consensus sequence across the two haplotypes of a diploid clone derived from a tetraploid elite variety and assess the sequence divergence from the available potato genome assemblies, as well as among the two haplotypes; (ii) to evaluate the new assembly’s usefulness for various genomic methods and (iii) to assess the performance of phasing in diploid and tetraploid clones, using linked read sequencing technology. We used PacBio long reads coupled with 10x Genomics reads and proximity-ligation scaffolding to create the dAg1_v1.0 reference genome sequence. With a final assembly size of 812 Mb, where 750 Mb are anchored to 12 chromosomes, our assembly is larger than other available potato reference sequences and high proportions of properly paired reads were observed for clones unrelated by pedigree to dAg1. Comparisons of the new dAg1_v1.0 sequence to other potato genome sequences point out the high divergence between the different potato varieties and illustrate the potential of using dAg1_v1.0 sequence in breeding applications.

Development of chromosome-specific markers for a study on introgressive hybridization of potato with the wild Mexican allotetraploid species Solanum stoloniferum Schltdl

In order to involve valuable germplasm of the wild Mexican allotetraploid potato species Solanum stoloniferum Schltdl. (genomic composition ААВВ) into breeding, pentaploid interspecific hybrids (ААAAВ) with cultivated potato S. tuberosum L. (АААА) and their backcross progenies are usually used. Homologous synapsis in meiosis of such hybrids is expected only between chromosomes of the A subgenome, therefore a question arose about a possibility of introgressing genetic material of the subgenome B into the A genome of cultivated potato. In this connection, development of various schemes for the B subgenome introgression into the genome of cultivated potato is considered as a topical issue. The previous research has yielded four schemes of S. stoloniferum involvement into breeding, which imply backcrossing with cultivated potato of the following interspecific hybrids: (1) hexaploids (genomic composition ААААВВ, the conventional introgression scheme), (2) tetraploids (putatively, АААВ), (3) self-pollination progeny of a 4x hybrid and (4) pentaploid hybrids with a putative genome composition of АААВВ. The present paper presents the first results of the development of chromosome-specific DNA markers for the identification of S. stoloniferum chromosomes in interspecific hybrids. An S. stoloniferum accession PI 205522 with a high degree of resistance to late blight and PVY had been found to possess several DNA-markers of the R-genes conferring resistance to these pathogens and was used in hybridization as a promising parent. A set of 23 SSR- and CAPS markers with the known chromosome location in S. tuberosum was generated. These markers detect polymorphism between parent genotypes, i.e., the diploid clone IGC 10/1.21 of cultivated potatoes S. tuberosum, and accession PI 205522 of S. stoloniferum. All the markers specific for the wild species were found in triploid (ААВ) and pentaploid (АААВВ) hybrids of S. stoloniferum × S. tuberosum. This set of markers will be used for efficiency assessment of different schemes for S. stoloniferum genetic material introgression into the obtained BC2-BC3 generations after crossing the interspecific hybrids with cultivated potato.

Application of Dna Cytometry In Complex Assessment of Individual Disease Prognosis

1. The results of DNA flow cytometry of prostate cancer samples indicate a large morpho-molecular heterogeneity of the tumor disease. In 40.8% of patients with breast cancer the tumor had a diploid clone and 59.2 – aneuploid, including tetraploid and other variants 2. Further study of the results of DNA cytometry and the accumulation of material in the comprehensive diagnosis of prostate cancer can shed light on the prognosis, clinical course and formation of therapeutic tactics of this pathology.

Fine Screening for Resistance to Cold-Induced Sweetening in Potato Hybrids ContainingSolanum raphanifoliumGermplasm

Potato is an indispensable part of human food. Many wild and cultivated potato relatives have been screened to find the best germplasm to improve productivity and quality, but only a small sample of the available biodiversity has been exploited. Most wild relatives are self-incompatible diploids. Genetic variability exists within and among populations, even within a species. Therefore, it is necessary to carry out fine screening to identify individuals carrying traits of interest. This study was carried out to quantify phenotypic variability for resistance to cold-induced sweetening, an important processing trait. Five families were evaluated for potato chip (crisp) color following cold storage of tubers harvested from four greenhouse trials and one field trial. The families were generated by crossing a single diploid clone to five plants from one accession of the wild potato relativeSolanum raphanifolium. Analysis of variance revealed that resistance against cold-induced sweetening was dependent on family and trial. This study underscores the importance of fine screening to select individuals in potato accessions for use in potato improvement.

Toward the development of highly homozygous diploid potato lines using the self-compatibility controlling Sli gene

Cultivated diploid potatoes (2n = 2x = 24) are self-incompatible, but can be altered to become self-compatible using the Sli gene. Previously, a diploid clone 97H32–6 was selfed up to S3 using the Sli gene. To explore the usefulness of the Sli gene for the production of highly homozygous diploid potatoes, 2 S4 families from the above 97H32-6 derived S3 lines (inbred series A) and 3 S5 families by continuous selfings from a different F1 (= S0) plant (inbred series B) were developed. The level of heterozygosity and the location of heterozygous loci on the genetic map were investigated using RFLP and AFLP markers. The average heterozygosity levels of the originally heterozygous loci decreased from 100% in S0 to 10.7% in S4 and 8.6% in S5 (inbred series A and B, respectively). The average rate of reduction in heterozygosity per generation (38.4% and 38.5% for inbred series A and B, respectively) was lower than the theoretically expected rate (50%). However, none of the loci or chromosome sections was exclusively heterozygous in the advanced self-progeny. Thus, highly homozygous and seed-propagated diploid potatoes could be obtained by repeated selfing using the Sli gene.Key words: Selfing, heterozygosity, diploid potato, Sli gene, Solanum.

The Potential for Green Fluorescent Protein as a Screening Tool in the Production of Haploid Potato

A hybrid between a highly regenerative diploid clone (BARD 1-3) of Solanum phureja and haploid inducer IVP 101 was transformed with Agrobacterium tumefaciens strain 4404 containing plasmid pHB2892 with genes for green florescent protein (GFP) and kanamycin resistance. Hemizygous primary transformants (To) were produced from three leaf discs: 17 diploid plants from one leaf disc, three and nine tetraploids from the other two leaf discs. GFP expression was observed qualitatively under fluorescence microscopes and quantitatively with a GFP meter. Segregation ratios for tetraploid T1 seedlings fit models for single duplex insertions (35 transgenic: 1 non) or double simplex insertions (15 transgenic: 1 non). Diploid T1 seedlings segregated for deleterious traits: dwarfed size and curled leaves, as well as the GFP transgene. Similar segregation patterns in diploid families implied that all diploids may have been from the same transformation event. The cumulative segregation showed the dwarfed and curled plants fit a single recessive gene ratio (3 normal: 1 mutant), and GFP fit a double-copy insertion ratio (15 transgenic: 1 non). Six T1 selections were free of deleterious traits, consistently high expressers of GFP, and produced fertile pollen.

Relationships between Floral Morphology, Breeding Behavior, and Flower Longevity in Easter Cactus

The effects of floral morphology and breeding behavior on flower longevity were investigated in Easter cactus [Hatiora ×graeseri (Werderm.) Barthlott)]. Four clones were studied: two diploid (n = 11) clones (`Evita' and `Purple Pride') that were highly self-incompatible (SI), and two cytochimeras (diploid epidermis and tetraploid subepidermis) that were recovered from the diploid cultivars, both of which were self-compatible (SC). The clones exhibited differences in the stage of floral development in which autogamy commenced. Autogamy commenced on the day of anthesis in the two `Evita' clones and occurred ≈5 days after anthesis in the `Purple Pride' cytochimera. In the `Purple Pride' diploid clone, anthers and stigmatic lobes remained spatially separated during the period from anthesis to senescence. Examination of styles collected from senesced, undisturbed flowers showed that few pollen tubes traversed to the base of the styles for the two SI diploid clones, whereas large numbers of pollen tubes were present at the base of the styles for the two SC cytochimeras. Flower longevity for the `Evita' cytochimera was significantly less than for `Evita' diploid, but the diploid and cytochimeral clones of `Purple Pride' exhibited similar flower longevities. Application of 2 mm silver thiosulfate, an inhibitor of ethylene (C2H4) action, did not affect flower longevity of `Evita' cytochimera. Our results show that flower longevity in Easter cactus is influenced by breeding behavior and the stage of floral development at which autogamy commences.