Somatic hybrids of Solanum tuberosum – application to genetics and breeding

Cultivated potato (Solanum tuberosum L.) is one of the first agricultural crops successfully cultured in vitro and used for obtaining of somatic hybrids. The review presents the current state of knowledge of somatic hybridisation involving this and other species from the genus of Solanum. Methods of somatic hybridisation, in particular factors that must be considered during designing the experiments are presented and discussed. The main attention however is focused on processes that are responsible for somatic hybrid formation. Complex interactions between genomes and plasmones lead to formation of symmetric, asymmetric and cytoplasmic recombinants. The concept of alloplasmic incompatibility is presented and discussed in relation to Solanum hybrids. Selected examples of potato somatic hybrids with agronomically important traits derived from wild species are presented in the table and discussed.     

Recombination in sesquidiploid hybrids of Lycopersicon esculentum × Solanum lycopersicoides and derivatives

Summary Sesquidiploid hybrids of L. esculentum (L) x S. lycopersicoides (S) were backcrossed to L via L. pennellii (P) as a bridging species in order to detect and measure recombination. Although use of P injected its traits into the populations, the investigated traits were proven to originate from S. The appearance of S traits in diploids in the immediate progeny of sesquidiploids but mainly of derived alien addition types proved the occurrence of recombination at rates varying from 1.6% to 16%. In subsequent BC's, these traits were inherited in dominant Mendelian fashion, except for deviations favoring recurrent parent alleles, sometimes with highly significant deviations from 11. Inheritance was investigated in BC and F₂ ex BC for 13 traits with strong phenotypic modifications of morphological, physiological, and isozymic nature. Monogenic determination was confirmed in most instances by tight linkages. For most of the traits, small progenies allowed only rough estimates of linkage intensities, but for Wa (gene for White anthers, universal in S), a test cross with four markers on chromosome 8 established its locus 2 cM distal to dl, proximally on 8L. Also noteworthy is the linkage of Dls, a gene determining sensitivity of flowering to long days, close to sp, situated subterminally on 6L. For the majority of traits, these manifestations of linkage proved that the appearance of S traits resulted from recombination, not alien chromosome substitution — a conclusion also reinforced by observations of chromosome pairing in alien addition types and diploid derivatives. Recombined S alleles have loci in various chromosome positions. Although they were discovered on the shorter chromosomes (nos. 6–12), hybridization barriers precluded tests with the longer chromosomes. Thus, no evidence was found for restriction of recombination to certain chromosomes or chromosomal regions. The prospects therefore appear favorable for deriving valuable traits from the S parent.     

Molecular cloning and biochemical characterization of α- and β-tubulin from potato plants (Solanum tuberosum L.)

Few studies have investigated microtubules from plants that host pathogenic fungi. Considerable efforts are underway to find an antimitotic agent against plant pathogens like Phytophthora infestans. However, screening the effects of antifungal agents on plant tubulin in vivo or using purified native microtubule in vitro is a time consuming process. A recombinant, correctly folded, microtubule-like structure forming tubulin could accelerate research in this area. In this study, we cloned full length cDNAs isolated from potato leaves using reverse-transcribed polymerase chain reaction (RT-PCR). Solanum tuberosum (Stub) α-tubulin and β-tubulin were predicted to encode 449 and 451 amino acid long proteins with molecular masses of 57 kDa and 60 kDa, respectively. Average yields of α- and β-tubulin were 2.0–3.5 mg l^?1 and 1.3–3.0 mg l^?1 of culture, respectively. The amino acids, His6, Glu198, and Phe170 involved in benomyl sensitivity were conserved in Stub tubulin. The dimerization of tubulin monomers was confirmed by western blot analysis. When combined under appropriate conditions, these recombinant α- and β-tubulins were capable of polymerizing into microtubules. Accessibility of cysteine residues of tubulin revealed that important ligand binding sites were folded correctly. This recombinant tubulin could serve as a control of phytotoxicity of selected antimitotic fungicide compounds during in vitro screening experiments.     

醋栗番茄Solanum pimpinellifolium遗传多样性分析

野生种醋栗番茄包含丰富的变异,具有众多优良性状。本研究对从世界不同地区收集的433份醋栗番茄遗传资源进行了表型鉴定及遗传多样性分析。表型鉴定结果表明,在收集的433份醋栗番茄中约有14%为樱桃番茄,370份典型醋栗番茄中约22%的材料存在不同程度的分离。群体变异分析表明,不同性状间变异系数存在较大差异,其中柱头变异系数最大,为56.716%;花瓣数遗传稳定,变异系数最小,为2.082%;相关分析表明,多个性状间存在显著相关性;利用表型和基因型数据聚类均将醋栗番茄群体划分为两大类群;主成分分析表明,坐果率、单果重、可溶性固形物对变异的贡献率较大。研究结果将为利用醋栗番茄进行栽培种番茄遗传改良奠定一定的基础。     

Asymmetric fusion between wild and cultivated species of potato (Solanum spp.) –detection of asymmetric hybrids and genome elimination

 The objective of this study was to evaluate the suitability of different techniques for a simple and rapid identification of asymmetric hybrids, without the use of selection markers and independent of the fusion partners used. Additionally, the degree of donor DNA elimination was determined. Among 473 viable plants obtained from asymmetric fusion experiments between three di-haploid breeding lines of potato (Solanum tuberosum) and diploid wild species (S. bulbocastanum, S. circaeifolium; X-ray treatment of the wild species) the most promising ones were investigated with three different methods: flow cytometry, RFLP analysis with an oligonucleotide probe (GATA)₄, and with single-copy probes. Flow cytometry, which combines a high screening capacity with detailed information about the DNA content and allows a distinction between asymmetric hybrids and chimeras, detected 31 hypo-tetraploid and 42 hypo-hexaploid regenerates among 224 plants. With the oligonucleotide probe (GATA)₄ only a few asymmetric hybrids were detected among all regenerates. More than 50% of these asymmetric regenerates were chimeras. Concerning the degree of DNA elimination, the results obtained by RFLP analysis with 17 single-copy probes were correlated with the results obtained by flow cytometry. The maximum DNA elimination of the donor genome was 52%. As a trend, an irradiation dosage of 210 Gy caused a higher DNA elimination in the wild species than a dosage of 70 Gy. No calli were obtained after irradiation of the wild species with 420 Gy. Received : 6 December 1996/Accepted: 24 January 1997     

Anther cultures of tetraploid Solanum genotypes—the influence of gelling agents and correlations between incubation temperature and pollen germination temperature

Four tetraploid potato genotypes (194.10, 199.13, 201.5, 201.12) were examined in anther culture. The androgenic responses were in general high. Cv. 199.13 contributed with the best response, varying between 0.38 and 0.55 embryoids per anther. Gellan gum or potato starch were used as gelling agents in a double-layer medium. Anthers incubated on potato starch gave a higher embryo yield in the beginning of the culture period, compared to anthers cultivated on gellan gum. The number of embryoids per anther, however, was higher on gellan gum at the end of the culture period. Anther cultures of potato were incubated in two different temperatures (20 °C and 25 °C), and the highest embryo yield was obtained in 25 °C except for genotype 201.12 where no difference was found between the two temperatures. Experiments with pollen germination in various temperatures (10 °C and 20 °C) were correlated to anther culture experiments. Also in case of pollen germination, genotype 201.12 was temperature-independent, while germination was stimulated by higher temperatures in the other genotypes.     

Characterization and expression profiling of selected microRNAs in tomato (Solanum lycopersicon) ‘Jiangshu14’

Presence of selected tomato (Solanum lycopersicon) microRNAs (sly-miRNAs) was validated and their expression profiles established in roots, stems, leaves, flowers and fruits of tomato variety Jiangshu14 by quantitative RT-PCR (qRT-PCR). In addition conservation characteristics these sly-miRNAs were analyzed and target genes predicted bioinformatically. Results indicate that some of these miRNAs are specific to tomato while most are conserved in other plant species. Predicted sly-miRNA targets genes were shown to be targeted by either by a single or more miRNAs and are involved in diverse processes in tomato plant growth and development. All the 36 miRNAs were present in the cDNA of mixed tissues and qRT-PCR revealed that some of these sly-miRNAs are ubiquitous in tomato while others have tissue-specific expression. The experimental validation and expression profiling as well target gene prediction of these miRNAs in tomato as done in this study can add to the knowledge on the important roles played by these sly-miRNAs in the growth and development, environmental stress tolerance as well as pest and disease resistance in tomatoes and related species. In addition these findings broaden the knowledge of small RNA-mediated regulation in S. lycopersicon. It is recommended that experimental validation of the target genes be done so as to give a much more comprehensive information package on these miRNAs in tomato and specifically in the selected variety.     

Remediation and Safe Production of cd Contaminated Soil Via Multiple Cropping Hyperaccumulator Solanum nigrum L. and Low Accumulation Chinese Cabbage

Multiple crop experiment of hyperaccumulator Solanum nigrum L. with low accumulation Chinese cabbage Fenyuanxin 3 were conducted in a cadmium (Cd) contaminated vegetable field. In the first round, the average removal rate of S. nigrum to Cd was about 10% without assisted phytoextraction reagent addition for the top soil (0–20 cm) with Cd concentration at 0.53–0.97 mg kg^?1 after its grew 90 days. As for assisted phytoextraction reagent added plots, efficiency of Cd remediation might reach at 20%. However, in the second round, Cd concentration in Chinese cabbage was edible, even in the plots with assisted phytoextraction reagent added. Thus, multiple cropping hyperaccumulator with low accumulation crop could normally remediate contaminated soil and produce crop (obtain economic benefit) in one year, which may be one practical pathway of phytoremediating heavy metal contaminated soil in the future.     

Nuclear and cytoplasmic genome components of Solanum tuberosum + S. chacoense somatic hybrids and three SSR alleles related to bacterial wilt resistance

The somatic hybrids were derived previously from protoplast fusion between Solanum tuberosum and S. chacoense to gain the bacterial wilt resistance from the wild species. The genome components analysis in the present research was to clarify the nuclear and cytoplasmic composition of the hybrids, to explore the molecular markers associated with the resistance, and provide information for better use of these hybrids in potato breeding. One hundred and eight nuclear SSR markers and five cytoplasmic specific primers polymorphic between the fusion parents were used to detect the genome components of 44 somatic hybrids. The bacterial wilt resistance was assessed thrice by inoculating the in vitro plants with a bacterial suspension of race 1. The disease index, relative disease index, and resistance level were assigned to each hybrid, which were further analyzed in relation to the molecular markers for elucidating the potential genetic base of the resistance. All of the 317 parental unique nuclear SSR alleles appeared in the somatic hybrids with some variations in the number of bands detected. Nearly 80 % of the hybrids randomly showed the chloroplast pattern of one parent, and most of the hybrids exhibited a fused mitochondrial DNA pattern. One hundred and nine specific SSR alleles of S. chacoense were analyzed for their relationship with the disease index of the hybrids, and three alleles were identified to be significantly associated with the resistance. Selection for the resistant SSR alleles of S. chacoense may increase the possibility of producing resistant pedigrees.     

Production and characterization of arboreous and fertile <Emphasis Type="Italic">Solanum melongena</Emphasis> + <Emphasis Type="Italic">Solanum marginatum </Emphasis>somatic hybrid plants

In crop plants the shift from being annuals to perennials may allow future agricultural systems requiring less energy inputs. The practicability of this was tested for Solanum melongena. Leaf protoplasts of S. melongena (2n = 2x = 24) and one of the related arborescent species Solanum marginatum (2n = 2x = 24) were electrofused and fertile somatic hybrids with arborescent habit regenerated. The magnetic cell sorter (MACS) technique was used for the selection of heterokaryons. The hybrid nature of 18 regenerated plants was assessed on the banding patterns generated by inter-simple sequence repeat PCR. When taken to maturity in the greenhouse, hybrids grew more vigorously compared to the parental species. Their morphological traits were intermediate between those of S. melongena and S. marginatum. Hybrids flowered and produced an average of 85% stainable viable pollen and fertile fruits. The somatic hybrids were maintained in the greenhouse for more than 3 years and continued to produce flowers developing into two types of fruits with plentiful seeds. Fruits were either striated green containing non-germinable seeds or yellow with fully germinable seeds. Their S₁ progenies showed common features with S₀ hybrids, including fertility and arborescent habit. Cytologically, somatic hybrids exhibited the expected chromosome number of 2n = 4x = 48, while chromosome pairing during microsporogenesis was associated with a low frequency of intergenomic pairing. It is concluded that an arborescent perennial species has been obtained by somatic hybridization. The usefulness of this species per se or in eggplant breeding will depend not only on the transmission of the arborescent habit to cultivated eggplant varieties, but also on the variability that should be created from backcrossing the S. melongena + S. marginatum hybrids to S. melongena.     

Mitigation of sodium chloride toxicity in Solanum lycopersicum L. by supplementation of jasmonic acid and nitric oxide

We investigated the effects of exogenous application of jasmonic acid (JA) and nitric oxide (NO) on growth, antioxidant metabolism, physio-biochemical attributes and metabolite accumulation, in tomato (Solanum lycopersicum L.) plants exposed to salt stress. Treating the plants with NaCl (200 mM) resulted in considerable growth inhibition in terms of biomass, relative water content, and chlorophyll content, all of which were significantly improved upon application of JA and NO under both normal and NaCl-stress treatments. Salt treatment particularly 200 mM NaCl caused an apparent increase in electrolyte leakage, lipid peroxidation, and hydrogen peroxide production, which were reduced by exogenous application of JA and NO. Salt treatment triggered the induction of antioxidant system by enhancing the activities of antioxidant enzymes like superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), and glutathione reductase (GR). Application of JA and NO separately as well as in combination caused a significant improvement in activities of SOD, CAT, APX, and GR activities. JA and NO either applied individually or in combination boosted the flavonoid, proline and glycine betaine synthesis under NaCl treatments. In conclusion, the exogenous application of JA and NO protected tomato plants from NaCl-induced damage by up-regulating the antioxidant metabolism, osmolyte synthesis, and metabolite accumulation.     

Solanum macolae,eine neue käferfeste Wildkartoffel

Theoretical and Applied Genetics -     

Assessment of direct shoot organogenesis and genetic fidelity in Solanum viarum Dunal—a commercially important medicinal plant

Solanum viarum Dunal is an important medicinal plant with a high quantity of steroidal alkaloids used for the synthesis of contraceptives, corticosteroids, and sex hormones. It is also used by Indian tribal people for the treatment of leprosy, toothache, and diabetes. Therefore, to meet the existing needs for this plant, it is necessary to develop an efficient regeneration system useful for rapid and large-scale clonal propagation with ensured genetic fidelity. An efficient and improved regeneration protocol for prickly and prickleless genotypes of S. viarum has been developed using three explants, leaf, petiole, and internodes, under the influence of two plant growth regulators, thidiazuron (TDZ) and 6-benzyladenine (BA). Effects of genotype, explant type, and concentrations of TDZ and BA were studied. A higher percentage of shoot organogenesis (78.25% ± 2.53) and shoot number per explant (6.79 ± 1.04) were achieved in the leaf segments of prickly genotype cultured on modified Murashige and Skoog (MS) medium supplemented with TDZ (1.50 mg L⁻¹). Furthermore, basal leaf segments showed 100% regeneration from the prickly genotype. A significantly higher content of total phenolics was quantified in prickleless (3.66 μg mg⁻¹) than prickly genotypes (2.73 μg mg⁻¹). The monomorphic banding pattern of random amplified polymorphic DNA (RAPD) and simple sequence repeats (SSR) analysis confirmed the genetic fidelity of the regenerated plants. Additionally, flow cytometric analysis of regenerants showed no variation in the ploidy levels when compared to the mother (control) plants. These results clearly depicted the efficiency of developed protocol that can be utilized for generating genetically stable population of S. viarum.

     

番茄(Solanum lycopersicum)类胡萝卜素降解关键基因筛选

类胡萝卜素衍生挥发物对提升番茄风味至关重要。为筛选调控类胡萝卜素衍生挥发物合成的关键基因,以90个番茄自交系中香气寡淡的TI4001和香气浓郁的CI1005为材料,分析了番茄类胡萝卜素裂解双加氧酶(SlCCDs)基因在不同组织及不同发育期果实中的表达量,果实不同成熟期类胡萝卜素及其衍生挥发物的含量。发现在7个SlCCDs基因中,SlCCD1A和SlCCD1B基因在番茄果实中表达量最高,且随着果实发育成熟表达量显著升高。果实中类胡萝卜素及其衍生挥发物含量也显著升高。SlCCD1A和SlCCD1B基因表达量与类胡萝卜素及其衍生挥发物含量之间极显著正相关。推测SlCCD1A和SlCCD1B基因是裂解类胡萝卜素合成挥发物的关键基因。     

Wild Solanum resistance to aphids: antixenosis or antibiosis?

The type (antixenosis or antibiosis) of resistance against the aphids Myzus persicae (Sulzer) and Macrosiphum euphorbiae (Thomas) was characterized for the wild tuber-bearing potatoes, Solanum chomatophilum Bitter and Solanum stoloniferum Schltdl. & Bouché through behavioral (olfactometry and electrical penetration graph) and physiological studies. In dual-choice assays, only S. stoloniferum exerted attraction to M. euphorbiae. This ruled out the possibility that plant volatiles of S. chomatophilum and S. stoloniferum may contribute to the high resistance expressed. In electrical penetration graph experiments, aphids feeding on S. stoloniferum showed increased salivation phases, whereas phloem ingestion was drastically reduced for both aphid species. Because reaching phloem elements was not delayed in both species, the resistance mechanism was phloem-located. The antixenosis exhibited by S. stoloniferum was similar on young and mature leaves. S. chomatophilum also showed phloem-located antixenosis against M. persicae. In contrast, M. euphorbiae had no difficulty to reach S. chomatophilum phloem tissues and to ingest sap. S. chomatophilum resistance against M. euphorbiae was antibiosis and only expressed in mature leaves, where a complete nymphal mortality was observed.     

The effects of IAA and tetcyclacis on tuberization in potato (Solanum tuberosum L.) shoot cultures in vitro

Potato (Solanum tuberosum cv. Désirée) shoots grown in vitro in continuous darkness or in long days (LDs), were used to investigate indole-3-acetic acid (IAA) effects on stolon initiation and tuber formation, combining IAA with increased or decreased gibberellin levels. An increased gibberellin (GA) level was achieved by the applying 1 μM GA₃, while decreased gibberellin level was presumably realized by the adding 3 μM tetcyclacis (Tc). About 15% of potato shoots developed stolons both in LDs and in darkness. Stolon initiation was stimulated by GA₃ in darkness and by Tc in LDs. Tuber formation was strongly inhibited in LDs and by GA₃ both in light and darkness, but stimulated in darkness at low GA level. Exceptionally, tuber formation occurred in LDs at the highest Tc concentrations, in about 25% of explants. Indole-3-acetic acid alone stimulated stolon formation in LDs, both in the presence or absence of GA₃. IAA alone also stimulated tuber formation in dark-grown shoots, but could not overcome the inhibitory effect of LDs. Indications that, depending on their concentration ratio, IAA may interact with GA₃ in different tuberization phases, have been discussed. Radomir Konjević—Deceased in July 2006.     

Does light spectral quality affect survival and regeneration of potato (Solanum tuberosum L.) shoot tips after cryopreservation?

Cryopreservation, the storage of germplasm at ultra-low temperature is the most reliable tool for long-term preservation of plant genetic resources. Cryopreservation techniques are widely applied but the effect of light spectra on plant recovery after cryopreservation is largely unknown. Therefore, we investigated the effect of different light spectral qualities on survival and regeneration of shoot tips of potato (Solanum tuberosum L.) cultivars Agrie Dzeltenie, Maret, Bintje, Désirée and Anti cryopreserved by the DMSO-droplet method. Prior to cryopreservation, the plants were stored under cool white fluorescent light (CW). Post-cryopreservation, the plants were allowed to regenerate under six different light qualities: CW, warm white light (HQI), blue LEDs (B), red LEDs (R), red with 10 % of blue (RB) and RBF - red with 10 % of blue with addition of 20 % of far-red LEDs. The light spectral quality had a significant effect on the survival and regeneration of potato shoot tips after cryopreservation. The combination of red light with 10 % of blue (RB) doubled the regeneration percentage of all cultivars, whereas red light (R) was not suitable for regeneration after cryopreservation. Specifically, the regeneration percentages were increased in RB compared to CW from 25.5 to 52.6 % for ‘Agrie Dzeltenie’, 25.0–43.6 % for ‘Maret’, 8.1–26.1 % for ‘Bintje’, 0.0–17.1 % for ‘Anti’ and 18.2–36.6 % for ‘Désirée‘. Therefore, the modification of light spectra during the recovery phase is a promising tool for increasing the regeneration of potato shoot tips after cryopreservation.     

Complex genetics controls natural variation among seed quality phenotypes in a recombinant inbred population of an interspecific cross between Solanum lycopersicum × Solanum pimpinellifolium

Seed quality in tomato is associated with many complex physiological and genetic traits. While plant processes are frequently controlled by the action of small‐ to large‐effect genes that follow classic Mendelian inheritance, our study suggests that seed quality is primarily quantitative and genetically complex. Using a recombinant inbred line population of Solanum lycopersicum × Solanum pimpinellifolium, we identified quantitative trait loci (QTLs) influencing seed quality phenotypes under non‐stress, as well as salt, osmotic, cold, high‐temperature and oxidative stress conditions. In total, 42 seed quality traits were analysed and 120 QTLs were identified for germination traits under different conditions. Significant phenotypic correlations were observed between germination traits under optimal conditions, as well as under different stress conditions. In conclusion, one or more QTLs were identified for each trait with some of these QTLs co‐locating. Co‐location of QTLs for different traits can be an indication that a locus has pleiotropic effects on multiple traits due to a common mechanistic basis. However, several QTLs also dissected seed quality in its separate components, suggesting different physiological mechanisms and signalling pathways for different seed quality attributes.