Parthenogenetic monohaploids (2n=x=12) from Solanum tuberosum L. and S. verrucosum Schlechtd. and the production of homozygous potato diploids

Summary Dihaploid and dihaploid derived clones of Solanum tuberosum and diploid genotypes of S. verrucosum produced 85 viable monohaploids by female parthenogenesis. All were induced using diploid S. phureja clones, homozygous for embryo spot, as pollinator. Frequency of S. tuberosum monohaploids per 100 berries was rather constant in three successive years (14, 17 and 17 respectively). No male and female fertility was found in flowering monohaploids.Colchicine-induced chromosome doubling yielded homozygous s. tuberosum diploids with low pollen quality but good seed fertility.Two diploid self-incompatible species (S. multidissectum and S. berthaultii) produced no monohaploids. The presence of genes for female parthenogenesis in some dihaploids is discussed.     

Monohaploid plants from anthers of a dihaploid genotype of Solanum tuberosum L.

Summary Monohaploid Solanum tuberosum plants were produced from the anthers of a dihaploid genotype. From another dihaploid genotype plants containing 36 chromosomes were obtained. For plantlet production anthers containing pollen at the uninucleate microspore stage were inoculated on a Linsmaier and Skoog-based medium supplemented with 1 mg/1 indole-3-acetic acid and 1 mg/1 benzyl aminopurine.Anthers from donor plants grown either in the summer or in the winter responded similarly on a range of media. Anther response in the form of callus induction and root formation was at least partly dependent on the genotype of the donor dihaploid plant.     

In vitro induction of tetraploid plants from colchicine-treated diploid daylily callus

Summary Daylily plantlets with tetraploid or octaploid chromosome numbers were induced from colchicine-treated diploid callus (2n=22). Callus tissue of the daylily Hemerocallis flava L. was placed on a morphogenetic induction medium, a modified MS formula supplemented with 1 mg 2,4-D and 1 mg kinetin/liter, plus 0,10, 20, or 40 mg/liter colchicine in the dark at 12°C for 3 days and recuperated for 1 week under the same environmental conditions except that colchicine was left out. The calluses were then returned to the normal growth temperature 25°C. Ploidy was identified by chromosome counts of squashed root-tip cells collected from the newly potted plantlets and later by measurements of stomates and pollen grains. Over 50% of the plants initiated from the colchicine-treated calluses were completely tetraploid. All of the plantlets differentiated from untreated callus had a diploid number of 22. Of the 3 colchicine levels applied, the 20 mg/liter treatment appeared to be the most effective in production of tetraploidy.     

Comparison of diploid and tetraploid potato families derived from Solanum phureja x dihaploid S. tuberosum hybrids and their vegetatively doubled counterparts

Summary To determine whether in potatoes the tetraploid level is preferable to the diploid level, especially regarding tuber yield, four diploid (2n=2×=24) Solanum phureja x dihaploid S. tuberosum hybrid parents and their vegetatively doubled, tetraploid (2n=4×=48) counterparts were intermated, which resulted in two F₁ hybrid families at both levels of ploidy. The parents and clones of the F₁ families and their offspring were used in crosses in such a way that in addition Sib₁, Sib₂, F₁×Sib₁, BC₁ and Sib₁×Sib₁ families were produced. Of the first clonal generation of the 12 2 x families and their 12 counterpart 4 x families two tubers per clone were planted in three replications in a field experiment at Sturgeon Bay in 1969; of the parents six tubers were planted in each replication. Data were recorded on 16 characters, including plant height at four different times.The ANOVA's showed significant clone effects within each family for all characters. Computed from all family means as well as from the family means per ploidy level, differences due to family were also significant for all characters except one.As at the 2 x level and at the 4 x level of ploidy the mean phenotypic correlations between characters were of similar magnitude, it is concluded that they are independent of ploidy level.With the exception of eye depth, the mean coefficient of variation was greater at the 2 x than at the 4 x level of ploidy, indicating the greatest response to selection for those characters at the 2 x level. From the differences in family means between the 4 x and the 2 x level of ploidy it was apparent that the 4 x families generally had significantly taller plants, later maturity, fewer tubers, higher mean tuber weight, more tuber yield and more dry matter yield than their counterpart 2 x families. In contrast, the 4 x parents had on the average shorter plants, lower mean tuber weight, much lower tuber yield and lower dry matter yield than their 2 x counterparts.The phenotypic correlation and Spearman's rank correlation between the family means of the 2 x and the 4 x level of ploidy were positive for almost all characters and significant for nearly half of them.From the results it is concluded that 1. in potatoes the 4 x level of ploidy is preferable to the 2 x level, and 2. the performance of 4 x families is predictable from the performance of their counterpart 2 x families.Based on results mentioned in the literature and on the present results, a continued use of S. tuberosum dihaploids in potato breeding needs to be dissuaded.     

A biosystematic study of the origin of the cultivated diploid potato,Solanum x Ajanhuiri Juz. et Buk.

Summary The origin and nature of the diploid cultivated potato species S. ajanhuiri Juz. et Buk. was studied. Several lines of evidence indicate that S. ajanhuiri might be derived from natural crosses between primitive cultivars of the diploid species S. stenotomum and the wild species S. megistacrolobum. Morphological comparisons were made between S. stenotomum x S. megistacrolobum F₁ hybrids and naturally occurring S. ajanhuiri to investigate this hypothesis. Comparisons were also made between S. ajanhuiri x S. stenotomum crosses and the F₂ generation of the first-mentioned cross.Crosses between the two major groups of S. ajanhuiri cultivars, Ajawiri and Yari, showed not only genetic breakdown but also a wide range of phenotypic variation similar to those of artificial F₂ families of S. stenotomum x S. megistacrolobum. Furthermore, there was strong evidence showing that the Yari group of S. ajanhuiri could almost certainly be an F₁ S. stenotomum x S. megistacrolobum hybrid, whereas the Ajawiri group could be a backcross of an F₁ hybrid to S. stenotomum. These results added further support to the hypothesis of a hybrid origin of S. ajanhuiri, as well as indicating its putative parents. It is suggested that this hybridogenic taxon be retained at the species level under the name Solanum x ajanhuiri.     

Isolation and characterisation of somatic hybrids of diploid Solanum tuberosum and Solanum brevidens and the use of amylose-free starch mutation for detection of introgression

Summary Somatic hybrids of diploid amylose-free (amf) Solanum tuberosum and diploid S. brevidens were made by Poly-Ethylene-Glycol (PEG) or electrofusion methods. For the isolation of interspecific hybrids the use of selection markers (kanamycin and hygromycin resistance) was useful but not essential. In this 2x+2x interspecific combination 4x and 6x somatic hybrids were obtained. Seed set was the best in 4x×4x (S. tuberosum) backcrosses, but seed germination was the best in 6x×4x combinations, using in vitro germination of unripe seeds harvested 25 days after pollination. A high degree of pollen stainability (30–40%) was observed in 7 tetraploid hybrids and very low in all hexaploids. After iodine staining, the recessive amf marker was expressed by a red colour instead of blue, visible in starch-containing cells like columella cells of root tips, (micro)tubers or microspores. As expected, complementation was observed in starch-containing cells of the fusion hybrids. Segregation of the amf marker was clearly observed in microspores of 4x and 6x hybrids. Segregation ratios in the 4x hybrids showed variable recombination frequencies. In the backcross progeny of hexaploid F12-5 with a tetraploid amf mutant one amylose-free recombinant among 67 plants was found, indicating the occurrence of meiotic recombination in the megaspore mother cells.     

Enhancement of diploid Solanum chacoense Bitt. using adapted clones of Solanum phureja Juz. & Buk.

Summary F₁ hybrids between Solanum phureja and S. chacoense were studied in a new approach to enhance wild diploid potato species using adapted S. phureja clones as the female parent. S. chacoense parents had few tubers, whereas S. phureja parents had been previously selected for tuberization under long days. Twenty hybrid families were evaluated for haulm vigor, vine maturity, stolon development, frequency of tuberization, tuber set, tuber yield and average tuber weight under naturally occurring increasing or decreasing photoperiods in spring and fall growing seasons in Morocco. The hybrid plants were more vigorous, later maturing with thicker stolons in the spring season. Frequency of tuberization and tuber set did not vary with season. Tuber yield and average tuber weight were higher in fall; the maximum yield was 2.4 and 4.6 kg per hill under spring and fall conditions, respectively. Remarkable variation was found both between and within hybrid families for all traits in this first cycle of enhancement. The selection of vigorous, fertile clones incorporating the S. chacoense genome is possible by using S. phureja as an adapted germplasm source.     

Plant regeneration from callus culture in potato

Summary A procedure for plant regeneration from callus culture of potato, Solanum tuberosum L. is described. Calli were induced from 1–2 mm long shoot apices of potato cultivars Cara and A25/19 on half-strength Murashige and Skoog's medium (half-MS) supplemented with 3.2 mg IAA (indole-3-acetic acid), 1.0 mg kinetin (6-furfurylamino)purine], and 0.5 mg 2,4-D [2,4-dichlorophenoxy)acetic acid]/1. Sixty percent explants produced nodular calli on this medium within 30 days. Calli differentiated into shoot-primordia when subcultured on half-MS medium supplemented with 0.5 mg 2,4-D and 1.0 mg zeatin [6-(4-hydroxy-3-methybut-2 enylamino)amino purine]/1. Differentiated calli on half-MS medium without growth hormones produced complete plantlets which were cloned on the same medium and transferred into soil.     

Occurrence of self-compatibility, self-incompatibility and unilateral incompatibility after crossing diploid S. tuberosum (SI) with S. verrucosum (SC): I. Expression and inheritance of self-compatibility

Diploid Solanum tuberosum (tbr), 2n=2x=24,can be crossed with S. verrucosum (ver) only when the latter is used as a pistillate parent but not reciprocally. This conforms to the phenomenon of unilateral incompatibility (UI) where a self-compatible species, like ver (SC) cannot be used as a male parent to cross with a self-incompatible (SI) parent like tbr. Even if ver × tbr hybrids are made, the F1 hybrids possess cytoplasmic male sterility and thus hinder genetic analysis of crossing barriers. Exceptionally, however, some diploid genotypes of tbr (SI) can be used as pistillate parents to cross with ver, and such exceptional tbr clones are called `acceptors'. Repeated backcrossing of acceptors to ver have resulted in male fertile genotypes that possess tbr cytoplasm and ver nucleus. These genotypes were used for the genetic analysis of `acceptance' and UI in thse experiments. It was found that acceptance of ver-pollen by tbr-pistils is based on a dominant gene A that expresses only in the absence of an inhibitor I. In the F₁ hybrids, only the S-allele of tbr was expressedbut not that of ver. Concomitant with this observation, it was shown that ver does not produce style-specific S-glycoproteins that are responsible for self-incompatible reaction in diploid potato. Although the the F₁ populations were SC, they segregated into SC and SI genotypes giving skewed segregation ratios for this trait. Because of this as well as the disappearance and re-appearance of SC trait in the offspring generations, it was necessary to postulate a more complex interaction between A and I. Models are presented in order to explain acceptance, non-acceptance and the expression of UI. It is concluded that at least four different loci are involved in the expression of UI. This revised version was published online in July 2006 with corrections to the Cover Date.     

The partial stability of additional chromosomes in Solanum tuberosum cv. Torridon

Summary Commercial cultivars of the potato (Solanum tuberosum L.) are widely believed to be cytologically invariable (2n=48), although there are several reports of anomalous counts in experimental material. The presence of two additional chromosomes is reported for the first time in cv. Torridon and their stability in the growing plant examined. The additional chromosomes were found to be completely stable in the shoots but were progressively eliminated from the root tissue. Laggards and bridges were commonly observed during somatic division in the roots. Laggards were also present in the shoots and did not change significantly in frequency in the roots during chromosomal loss. Consequently, they were considered as unlikely to have a central involvement in the elimination process. Somatic bridges on the other hand were only found in the roots and increased in occurrence as elimination increased. A link between the appearance of somatic bridges and the elimination of additional chromosomes in the roots is suggested. The possible origins and mechanism of elimination of the additional chromosomes are briefly discussed.     

Regeneration of plants from potato meristems freeze-preserved for 24 months

Summary Plants have been regenerated from cultures of potato meristems freeze-preserved in liquid nitrogen (–196°C) for 24 months. The implications and the feasibility of employing cryogenic methods for the long-term conservation, and international exchange of germplasm of potato, and other vegetatively propagated crops are pointed out.     

An assessment of morphogenic and transformation efficiency in a range of varieties of potato (Solanum tuberosum L.)

Summary To provide a truly genotype-independent transformation system, it is necessary to be able to transform a wide range of potato genotypes. The ability to regenerate shoots in vitro was determined for 34 potato varieties using tuber disc explants. Following a culture regime used extensively in previous studies with the variety Desiree, half of the varieties could be regenerated from tuber discs and half could not. From a sample of varieties that could be regenerated from tuber discs, all but one variety gave transgenic plants. Twelve varieties were evaluated for the capacity to regenerate shoots from leaf and internode explants excised from in vitro grown plants. All of the varieties tested regenerated adventitious shoots. Leaf and internode explants from 5 varieties were subsequently used for transformation, and transgenic plants were produced from two potato varieties that did not give transgenic plants from tuber disc explants. Some varieties could not be transformed by either method, and will require modification of the in vitro regeneration and transformation system to be successful.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - GA₃ gibberellic acid - IAA indole-3-acetic acid     

Molecular stability of the ribosomal RNA genes in Solanum tuberosum plants recovered from slow growth and cryopreservation

Summary Ribosomal gene (rDNA) probes have been used to assess genomic changes in plants of (1) S. tuberosum cv Desiree subjected to slow growth and (2) S. tuberosum cv Golden Wonder recovered from cryopreservation. Restriction fragment length polymorphisms (RFLPs) were detected in two out of 16 DNA samples extracted from plants derived from slow growth, control plants did not show RFLP differences. Plants recovered from cryopreserved shoot-tips of Golden Wonder were unchanged in their ribosomal gene RFLP profile compared to the untreated controls. The use of rDNA probes as tools to assess stability, and the possible detrimental effects of slow growth, somaclonal variation, cryoprotectants, and freezing injury are discussed.     

Reciprocal differences between hybrids of Solanum tuberosum Groups Tuberosum (haploid) and Phureja

Summary Yield components including total tuber yield, total tuber number, vine maturity, vine vigor and average tuber weight as well as seed germination and pollen stainability were studied in 28 exact reciprocal hybrid families, between 12 Solanum tuberosum Group Tuberosum (haploid) (TH) and 12 S. tuberosum Group Phureja (PH) clones in a NC Mating Design II in 4 sets of 3 by 3 reciprocal hybrids at three locations, one in Minnesota (USA) and two in Morocco. TH parents were insensitive to daylength whereas most PH clones required critical photoperiods equal to or less than 14 hours for tuberization. Mean reciprocal differences were significant for most traits at all 3 locations. Hybrids with TH cytoplasm were superior to those with PH cytoplasm by 18% for tuber yield, 21% for tuber number, and 9% for vine maturity, but inferior to hybrids with PH cytoplasm by 11% for vine vigor and 19% for time to reach 80% seed germination. Reciprocal differences were not detected for average tuber weight. Six reciprocal crosses revealed significant differences for pollen stainability but the overall mean difference (one location) was not significant. Possible reasons for the observed reciprocal differences included gamete selection, pseudo-self-compatibility and/or interaction of cytoplasm with both nuclear genes and environment. Differences for critical photoperiod of TH and PH parents were also demonstrated as a probable source of reciprocal differences.Scientific Journal Series Article 14896 of the Minnesota Agricultural Experiment Station supported in part by the U.S. Agency for International Development, contract no. 608-0160, IAV Morocco Project.     

Identification of the trisomic series in diploid Solanum tuberosum L. group Tuberosum. II. Trivalent configurations at pachytene stage

Summary By means of pachytene analysis 11 of the 12 possible types of primary trisomics of diploid S. tuberosum have been identified. The typical trivalent configurations for each chromosome are illustrated. Triplo-4,5,7 and 9 occurred more frequently whereas triplo-1 is still lacking in the series. Some of the features of trivalent associations at pachytene are discussed, as also the fact that in the present study only primary trisomics have been found.     

Interspecific hybridization between Solanum melongena and Solanum macrocarpon: study of the F1 hybrid plants

Summary Egg plant is one of the most common vegetable crop grown in India and other parts of the world. The cultivated egg plant Solanum melongena is found to be susceptible to the shoot and fruit borer (Leucinodes orbonalis). Whereas the species Solanum macrocarpon is resistant. In order to incorporate the resistance of shoot and fruit borer to the cultivated egg plant, an interspecific hybridization between Solanum melongena and Solanum macrocarpon and reciprocals were carried out. The hybrids were found to be sterile, the investigations revealed that the failure of seed set in hybrids is due to the ovule abortion. In order to overcome the sterility, the colchicine has been applied to the interspecific hybrids.     

Genetics of self-compatibility in dihaploids of Solanum tuberosum L. 5. The S-genotype of tetraploid potato cultivar gineke

Summary Investigations of the genetics of self-compatibility and self-incompatibility in dihaploids and diploid derivatives from cv. Gineke revealed the presence of S ₁, S₂ and S ₃ at the S-locus of Gineke and in addition an S ₁-allele on a translocation. By means of a complete tester set involving the S-alleles S ₁, S₂ and S ₃ (all from Gineke) and S ₄ (from Black 4495) it was demonstrated that some Gineke dihaploids were compatible with all six testers. This indicated a fourth S-allele in Gineke, which differs from those in the tester series and was therefore assigned S ₅. Additional evidence was obtained from an analysis of F₁'s from crosses of two S ₅-bearing dihaploids and one of the testers. So the S-genotype of cv. Gineke was identified as S ₁S₂S₃S₅/S₁, the second S ₁ being the S-allele on a translocated fragment.     

A comparative assessment of DNA fingerprinting techniques (RAPD, ISSR, AFLP and SSR) in tetraploid potato (Solanum tuberosum L.) germplasm

Several DNA marker systems and associated techniques are available today for fingerprinting plant germplasm but information on their relative usefulness in particular crops is limited. The study investigated PCR based DNA fingerprinting in a set of 39 potato cultivars using RAPDs (20 primers), ISSRs (6 primers), AFLPs (2 primers) and SSRs (5 primer pairs). Results show that each of the four techniques can on their own, individually identify each cultivar, but that techniques differ in the mean number of profiles generated per primer (or primer pair) per cultivar, referred to as Genotype Index (GI). The order of merit based on this criterium and in this material was AFLPs (GI = 1.0), a multi-locus SSR (GI = 0.77),RAPDs (GI = 0.53), ISSRs (GI = 0.47) and single locus SSRs (GI = 0.36). Problems in relating banding patterns to individual loci and alleles for polyploid genomes, using these techniques as they are currently employed, are also discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Efficient production of doubled haploid Brassica napus plants by colchicine treatment of microspores

Summary The effect of colchicine on isolated microspore cultures of Brassica napus was evaluated in order to combine a positive effect of colchicine on the induction of embryogenesis with the possibility to induce chromosome doubling at an early developmental stage, thus avoiding the production of haploid or chimeric plants. Colchicine was added to the culture medium immediately after isolation of B. napus microspores. The cultures were incubated from 6 to 72 h with various concentrations of colchicine. Samples were taken from the regenerating embryoids after 6 weeks for ploidy determination by flow-cytometry.The highest diploidization rate was obtained after a 24 h treatment of microspores with 50 mg/l colchicine, leading to 80–90% diploid embroids. A concentration of 100 mg/l colchicine applied for the same duration resulted in a lower diploidization rate (76–80%). Treatment durations of 6 h were not long enough to induce a high rate of diploidization, whereas the application of 10 mg/l for 72 h was also very effective.A sample of the plants regenerated from the colchicine treated microspores was transferred to the greenhouse. The plants looked similar to normal diploid rapeseed plants and showed reasonable pod and seed set. Thus, an additional generation for seed increase in the greenhouse is rendered unnecessary. The advantage of applying a minimum volume of colchicine under controlled in vitro conditions means a considerable saving of time and labour in DH-breeding programs.     

Female fertility of potato (Solanum tuberosum ssp. tuberosum) dihaploids

Summary Results of testcrosses and seed set data of the dihaploid breeding program at the Potato Research Institute in Gross Lüsewitz, Germany, indicate that the low level of female fertility could hinder success in crossing work. With the objective to reveal major components of female fertility, the behavior of 47 outstanding potato dihaploids (S. tuberosum ssp. tuberosum, 2n=2x=24) was assessed in testcrosses over a period of 3 years. Analysis of variance was carried out for the traits: number of seed per berry (S/B), number of intact seed per berry (IS/B), and number of non-germinable seed per berry (NGS/B). Significant sources of variation included cross combinations, years, and cross combination-year interactions. More than 20 S/B were obtained in 25.4% of all cross combinations performed over 3 years using up to three pollinators. Berry set was not correlated to seed set, but was in general determined by the pollinator used. The number of seedless berries (SLB/F) was related to neither total berry set nor seed production. The trait S/B remained stable over different years, and thus it deemed to be most useful as a measure of the level of female fertility in practical breeding work. Selection for female fertility in early generations of a breeding program is recommended to avoid limitations in the production of variability via sexual recombination.