Somatic hybridization as a tool for tomato breeding

Protoplast fusion can be used to produce somatic hybrids of species that cannot be obtained by sexual hybridization. The possibility to introgress genes from Solanum species into the cultivated tomato species Lycopersicon esculentum, and to obtain novel cytoplasm-nucleus combinations (cybrids) was considered as an important strategy to extend the genetic variation available for tomato breeding. Somatic hybrids between L. esculentum and other Lycopersicon species, as well as between L. esculentum and Solanum or Nicotiana species, have been produced. Specific mutants, genotypes with antibiotic resistances, and metabolic inhibition by iodoacetate or iodoacetamide and irradiation were used for the selection of hybrids. In addition, the improvement of protoplast culture techniques and the use of the favourable tissue culture traits derived from species such as L. peruvianum, which have been introduced into tomato by classical breeding, allowed the efficient recovery of somatic hybrids. However, the occurrence of somatic incongruity in fusion combinations of L. esculentum and Solanum and even more in L. esculentum and Nicotiana, did not allow the production of true cybrids and/or fertile hybrids, indicating the importance of both cytoplasm-nucleus and nucleus-nucleus interactions in somatic incongruity. Another problem with fusions between distantly related species is the strongly reduced fertility of the hybrids and the very limited homoeologous recombination between chromosomes of the parental species. Partial genome transfer from donor to recipient through microprotoplast (+) protoplast fusion, and the production of monosomic or disomic chromosome addition lines, light overcome some of these problems. In symmetric somatic hybrids between L. esculentum and S. tuberosum the occurrence of limited somatic and meiotic recombination was demonstrated. Fertile progeny plants could be obtained, though at a low frequency, when embryo rescue was performed on a large scale after backcrossing hexaploid somatic tomato (+) potato hybrids with a tetraploid potato genotype. The potential value of genomic in situ hybridization (GISH) and RFLPs for the analysis of the genome/chromosome composition of the hybrids has been demonstrated for intergeneric somatic hybrids between Lycopersicon and Solanum.Abbreviations cpDNA chloroplast DNA - mtDNA mitochondrial DNA     

Somatic Fusion for Breeding of Tetraploid Potatoes

From two tetraploid, one Transformed tetraploid, one triploid and 11 dihaploid clones of Solanum tuberosum somatic hybrids were produced by polyethylene glycol mediated somatic fusion. The inter-dihaploid clones comprised clones of agronomic value, homozygous doubled monohaploids, and in vitro selected clones resistant t0 Fusarium or Phytophthora toxins. Presumptive hybrids were enriched at the callus Stage in vitro by using differentiating media and by growth characteristics; further identification was performed by chromosome counting in vitro shoots and by isozyme analysis of in vitro plants. Final analysis was made from morphological characteristic of plant and tuber phenotypes. From 15 different combinations, 6009 plantlets have been regenerated. From five combinations, 310 reentrants were checked for hybrid nature by morphology and cytology and 88 by peroxidase and esterase isozyme analyses. Amongst these, from two combinations, a total of 17 different hybrids were confirmed by all methods. The procedures described are general enough to allow genome combination of interdihaploids resulting in tetraploids of practical breeding value.     

Monohaploids (n=x=12) from autotetraploid Solanum tuberosum (2n=4x=48) through two successive cycles of female parthenogenesis

Summary Viable monohaploids with 12 chromosomes in their somatic cells were obtained from an autotetraploid cultivar of Solanum tuberosum (2n=4x=48) by inducing female parthenogenesis successively in the tetraploid cultivar and in dihaploids from that cultivar. Both dihaploids and monohaploids were induced using the diploid S. phureja clones IvP 35 and IvP 48 as pollinators. The average frequencies of dihaploids and monohaploids in 1973 were 2.0 and 0.14 per berry respectively.Non-homologous chromosome associations (bivalents, trivalents and even quadrivalents) were observed at metaphase I of meiosis in pollen mother cells of the two monohaploids studied.The occurrence of non-homologous associations of chromosomes during meiosis is discussed. In addition the potential significance of monohaploids for basic research andfor breeding of potatoes is considered.     

Exploitation of the incompatibility mechanism for the production of F1 hybrid forage grasses

Summary The utility of the two locus (SZ) incompatibility system for the production of F₁ hybrids in forage grasses is examined in relation to its mode of action at the tetraploid level and in dihaploids. Schemes are proposed for producing tetraploid populations with a high level of within population incompatibility or of single or double cross hybrids from the dihaploids.     

Characterization of morphological variation and cold resistance in interspecific somatic hybrids between potato (Solanum tuberosum L.) and S. brevidens Phil.

Summary Somatic hybrids between Solanum tuberosum L. cv. Gracia (2n=4x=48) and Solanum brevidens Phil. (2n=2x=24) were produced via fusion of mesophyll protoplasts. Selection of the protoplast derived putative hybrid calli was based on their vigorous growth. Additive isozyme patterns and chromosome numbers as well as the expression of parental morphological characters have proved the hybrid origin of the selected regenerants. Extensive chromosome loss during the regeneration process resulted in aneuploid hybrids with high frequency. Genomic instability could not be detected in these plants during the period of vegetative propagation. Regenerants from hybrid tissues exhibited wide morphological variation especially in tuber formation. The detailed morphological analysis based on the use of multivariate method (principal component analysis, PCA) enabled to identify morphological groups among the hybrid clones. The positioning of hybrid clones in the PCA space could not be correlated with chromosome numbers. The genomic ratio represented by the tetraploid and diploid parents influenced the morphology of somatic hybrid population according to the applied analytical system. Two selected hybrid clones have exhibited an intermediate degree of frost tolerance compared to the parents, based on the recovery of plants from lower buds after freezing of potted plants.     

Polyploidy and sexual polyploidization in the genus Vaccinium

Genus Vaccinium,consisting of blueberries, cranberries, lingonberries and many related wild species, includes diploid, tetraploid, and hexaploid species. Most evidence indicates that the tetraploid species are autotetraploids, with non-preferential bivalent chromosome pairing. Although homoploid interspecific crosses usually produce numerous fertile hybrids if the parents are from the same section of the genus, inter sectional crosses at the diploid level normally produce no seedlings, weak seedlings, or seedlings that are very low in fertility. There is a strong but not complete triploid block within Vaccinium. Even with insections, tetraploid × diploid (and the reciprocal) crosses normally give only a few tetraploid and a few triploid hybrids. Hexaploid × diploid crosses within sections are very hard to make, and the few hybrids that have been obtained are pentaploid. The frequency of 2n gametes varies,both among genotypes within species and among species. Vaccinium pollen is normally shed in tetrads, and the frequency of large pollen grains shed in dyads can be used to estimate 2n gamete frequency. Cultivated blueberries occur at both the tetraploid and the hexaploid levels, and there are important genetic resources in the diploids. Unreduced gamete production has permitted transfer of genetic material from the diploid to the tetraploid level and from the diploids and tetraploids to the hexaploid level via triploid hybrids. Intersectional crosses can occasionally produce tetraploid Vaccinium hybrids that appear to behave as amphidiploids and have medium to high fertility. CommercialVaccinium cultivars are normally propagated by cuttings. Intersectional hybridization, chromosome doubling, and asexual propagation could permit the production of novel hybrid combinations with value as ornamentals or in fruit production. This revised version was published online in July 2006 with corrections to the Cover Date.     

Analysis of meiotic behavior in selecting potential genitors among diploid and artificially induced tetraploid accessions of Brachiaria ruziziensis (Poaceae)

Sexuality is correlated with diploidy and apomixis with polyploidy in the Brachiaria genus. Brachiaria ruziziensis is a key species in Brachiaria breeding due to its obligate sexuality and intrinsic agronomic qualities. Interspecific crosses in the genus became feasible only when a few diploid accessions of B. ruziziensis were artificially tetraploidized and remained sexual. Hybridization has been done since, using natural tetraploid apomictic accessions of B. brizantha or B. decumbens as pollen donors. Twenty two accessions of B. ruziziensis from the Embrapa Beef Cattle germplasm collection (Campo Grande, MS, Brazil) were cytologically analyzed: 16 are natural diploids (2n = 2x = 18) and six are artificially induced tetraploids (2n = 4x = 36). The meiotic behavior in the 16 diploid accessions varied. The mean of meiotic abnormalities per accession ranged from zero to 24.46%. Meiotic behavior in the induced tetraploid accessions also varied with the mean of meiotic abnormalities ranging from 5.20% to 54.71%. The most common abnormalities observed in both the diploid and the tetraploid accessions, were those related to irregular chromosome segregation. In one tetraploid accession, with a high frequency of those, other irregularities involving chromosome orientation at metaphase plate and chromosome convergence to the poles, a meiotic mutation known as divergent spindle, were recorded. Meiotic behavior should be considered in selecting potential genitors for breeding.     

Interspecific hybridization in <Emphasis Type="Italic">Sarcococca</Emphasis> supported by analysis of ploidy level,genome size and genetic relationships

Knowledge of ploidy level differences, genome size and genetic relationships between species facilitates interspecific hybridization in ornamentals. For Sarcococca (Buxaceae) only limited (cyto)genetic information is available. The aim of this study was to determine the genome size and chromosome number and to unravel the genetic relationships of a breeder’s collection using AFLP marker analysis. Based on these results, interspecific crosses were made and the efficiency and hybrid status was verified. Two groups of diploid plants (2n = 2x = 24) were observed, with either a genome size of 4.11–4.20 or 7.25–9.63 pg/2C. All the tetraploid genotypes (2n = 4x = 48) had genome sizes ranging from 7.91 to 8.18 pg/2C. In crosses between parents with equal ploidy level and genome size a higher crossing efficiency (on average 58% of the hybridizations resulting in fruits) and more true hybrids (on average 96% of the offspring) were obtained compared to crosses between plants with different genome size and ploidy level (on average 23% fruits and 24% hybrids, respectively). In none of the cross combinations, the ploidy level or genome size was found to be a complete hybridization barrier, although unilateral incongruity was found in some cross combinations. Distant genetic relationships did not hamper the hybridization within Sarcococca genotypes. Our findings will contribute to a more efficient breeding program and a faster achievement of hybrids with an added value.     

Double-bridge hybrids of Solanum bulbocastanum and cultivars of Solanum tuberosum

Summary Solanum bulbocastanum (2n=2x=24) has valuable characters for potato breeding, but cannot be hybridized directly with S. tuberosum cultivars. Both S. acaule (2n=4x) and S. phureja (2n=2x) were used as bridging species. Triploid S. acaule × S. bulbocastanum were doubled with colchicine and the resulting fertile hexaploid F₁'s crossed with S. phureja. The triple hybrids obtained were tetraploid or nearly so. The two genomes of S. acaule in these triple hybrids probably pair preferentially, which may provoke pairing and possibly crossing over between the chromosomes of S. bulbocastanum and S. phureja.More than 20000 pollinations of the triple hybrids with four potato cultivars had to be made to produce 40 quadruple hybrids. These highly vigorous hybrids varied greatly in many morphological characters, resistance to Phytophthora infestans, fertility and crossability. The chromosome numbers are 48 (24 hybrids), 49 and 46, but some higher ploidy levels (65, 66, 72 chromosomes) were found as well. Their origin is to be sought in the fusion of an unreduced egg cell from triple hybrids (either euploid or hypoploid) and a reduced male gamete from the cultivars. This view is corroborated by their extreme resistance to Phytophthora. Also some 48-chromosome hybrids are highly resistant, which may indicate introgression from S. bulbocastanum.Most quadruple hybrids are readily inter-crossable and crossable as females with cultivars; several also as males. Two could be hybridized with S. bulbocastanum, but the few seeds dit not germinate.Studies of pachytene stage of meiosis revealed the presence of a S. bulbocastanum chromosome in at least one tetraploid hybrid, which is highly resistant to Phytophthora. At metaphase I of meiosis chromosome associations higher than quadrivalents were not found. Except in one hybrid, the frequency of quadrivalents did not exceed one per cell and the average proportion of chromosomes associated as bivalents amounted to 90%.The quadruple hybrids (double-bridge hybrids) appear good starting material for breeding programmes aimed at introducing genes from S. bulbocastanum into S. tuberosum cultivars.     

In vitro induction of tetraploid plants from callus cultures of diploid bananas (<Emphasis Type="Italic">Musa acuminata</Emphasis>, AA group) ‘Kluai Leb Mu Nang’ and ‘Kluai Sa’

Tetraploid plants were induced successfully from diploid bananas Musa acuminata (AA genome) ‘Kluai Leb Mu Nang’ and ‘Kluai Sa’ (2n = 2x = 22) with in vitro oryzalin treatment. Calluses from in vitro-grown shoot tips of both cultivars were treated with oryzalin at concentrations of 1.5 or 3 mg l⁻¹ for 24, 48 and 72 h, respectively. The oryzalin treatments produced tetraploids at a frequency of 15.6% in ‘Kluai Leb Mu Nang’ and 16.7% in ‘Kluai Sa’ as detected by flow cytometry. Chromosome counting showed that the tetraploid plant chromosome number was (2n = 4x = 44). The selected tetraploid plants were transplanted in the field and variations in the morphological characteristic of leaf shape and fruit bunch compared to normal diploid plants were found under the same growing condition even after 3 years of cultivation.     

The use of dihaploids in increasing the homozygosity of tetraploid potatoes

Summary Electrophoresis banding patterns of the tuber proteins of 12 dihaploids of the cultivar Pentland Crown showed that four had types of patatin unlike that of the parent. The patatin types of somatically chromosome-doubled clones derived from three of the dihaploids were identical to those of the dihaploid progenitors. Fourteen dihaploids produced from the chromosome-doubled derivative (PDH40X2) of one dihaploid, which had a variant patatin, had the patatin type of the parent dihaploid.The experiment showed that dihaploids and somatic chromosome doubling could be used to fix variation found in heterozygous tetraploid potatoes. Used together, haploidisation and chromosome doubling can generate highly homozygous tetraploids for use by plant breeders.     

Alteration of the genomic composition of Solanum nigrum (+) potato backcross derivatives by somatic hybridization: selection of fusion hybrids by DNA measurements and GISH

Fusion experiments were performed with a first (BC₁‐6738) and a second (BC₂‐9017) generation backcross hybrid of 6x Solarium nigrum (+) 2x potato somatic hybrids with potato cultivars. Because no progeny was obtained from the BC₂ genotypes, alternative approaches were sought to overcome the sexual crossing barrier. Five potato genotypes, one of which contains the hygromycin resistance gene, were used in the fusion experiments. All vigorous regenerants were used for the estimation of nuclear DNA content using flow cytometry. Plants with a DNA content higher than that of the BC₁‐6738 or BC₂ genotypes were considered potential somatic hybrids. Forty‐nine potential somatic hybrids resulted from fusion experiments with BC₁‐6738, from which 20 grew vigorously in the greenhouse and flowered. After pollination with several 4x potato cultivars, eight genotypes produced seeded berries and five genotypes gave seedless berries. In addition, 11 of these 13 somatic hybrids were selected for genomic in situ hybridization (GISH) analysis to determine their genomic composition. Nine had exactly or approximately the expected number of 36 S. nigrum and 60 potato chromosomes. In one genotype, only 22 instead of 36 S. nigrum chromosomes were found and one potato chromosome was possibly missing. Only five potential somatic hybrids were detected among the 79 regenerants from BC₂‐9017 (+) 2x potato fusion experiments that were analysed by flow cytometry. Two of these hybrids were rather vigorous and did flower, but pollinations with potato have not yet set any berries.     

Progenies of allotriploids of Oriental × Asiatic lilies (Lilium) examined by GISH analysis

With the aim of utilizing allotriploid (2n = 3x = 36) lily hybrids (Lilium) in introgression breeding, different types of crosses were made. First, using diploid Asiatic lilies (2n = 2x = 24), reciprocal crosses (3x − 2x and 2x − 3x) were made with allotriploid hybrids (AOA) obtained by backcrosses of F₁ Oriental × Asiatic hybrids (OA) to Asiatic cultivars (A). Secondly, the AOA allotriploids were crossed with allotetraploid (OAOA, 2n = 4x = 48), in 3x − 4x combination. Finally, the AOA allotriploids where crossed to 2n gamete producer F₁ OA hybrids (3x − 2x (2n)). Two types of triploids were used as parents in the different types of crosses, derived from: (a) mitotic polyploidization and (b) sexual polyploidization. Ploidy level of the progeny was determined by estimating the DNA values through flowcytometry as well as chromosome counting. The aneuploid progeny plants from 3x − 2x and reciprocal crosses had approximate diploid levels and in 3x − 4x crosses and 3x − 2x (2n) the progeny had approximate tetraploid levels. Balanced euploid gametes (x, 2x and 3x) were formed in the AOA genotypes. Recombinant chromosomes were found in the progenies of all crosses, except in the case of 2x − 3x crosses through genomic in situ hybridization (GISH) analyses. Recombinant chromosomes occurred in the F₁ OA hybrid when the triploid AOA hybrid was derived through sexual polyploidization, but not through mitotic polyploidization with two exceptions. Those recombinant chromosomes were transmitted to the progenies in variable frequencies.     

Identification of somatic hybrids of dihaploid Solanum tuberosum lines and S. brevidens by species specific RAPD patterns and assessment of disease resistance of the hybrids

Summary Symmetric somatic hybrids were produced by electrofusion of protoplasts of two dihaploid tuber-bearing potato (Solanum tuberosum L.) lines and Solanum brevidens Phil., a diploid non-tuber-bearing wild potato species. A total of 985 plants was obtained. Verification of nuclear hybridity of putative hybrids was based on additive RAPD patterns, general morphological characteristics and chromosome counts. 53 (90%) calli regenerated into plants which were identified as somatic hybrids. Most of the hybrids were aneuploids at the tetraploid (4×) or hexaploid (6×) level. The 20 hybrids tested expressed a high level of resistance to potato virus Y (PVY ^N ) characteristic of the S. brevidens parent. Resistance to late blight (Phytophthora infestans (Mont.) de Bary) varied between hybrids, but was on average better than that of the fusion parents. Resistance of hybrids to bacterial stem rot (Erwinia carotovora subsp. atroseptica (van Hall) Dye) was not superior to that of commercial potato cultivars.     

Novel interspecific hybridization between sweetpotato (<Emphasis Type="Italic">Ipomoea batatas</Emphasis> (L.) Lam.) and its two diploid wild relatives

Interspecific hybridization is an important approach to broaden the genetic base and create novel plant forms in breeding programs. However, interspecific hybridization in Ipomoea is very difficult due to the cross incompatibility. Here we report two novel interspecific F₁ hybrids between I. batatas (L.) Lam. (2n = 6x = 90) and two wild species, I. grandifolia (2n = 2x = 30) and I. purpurea (2n = 2x = 30). Hybridization was stimulated by applying plant growth hormones. Morphological, molecular and cytological tests were conducted to confirm their hybridity. We found that the two hybrids were quite distinctive in leaf color and morphology, and yielded intermediate sizes of storage roots compared to their respective parents. Inter-simple sequence repeat analysis showed that the unique DNA bands from the wild parents could be detected in these two hybrids. The cluster analysis also showed that the two F₁ hybrids were closer to I. batatas in phylogeny relationship. The number of chromosomes of the two hybrids was both 60, indicating that the hybrids were tetraploid. The meiotic configuration analysis of the H₁ of I. batatas × I. grandifolia revealed the occurrence of 17.58I + 14.28II + 1.36III + 2.48IV at metaphase I in average chromosome association per pollen mother cells (PMCs), 4.26I + 18.32II + 2.56III + 3.12IV was average meiotic configuration in the H₂ of I. batatas × I. purpurea. Both hybrids appeared to be polyads and multi-microcytes at tetrad phase and differed in their pollen fertility.     

Inheritance of dwarfism in plantain (Musa spp., AAB group)

Plantain (Musa spp., AAB group) is a major food crop in the humid lowland tropics of Africa, the Caribbean and Latin America. Lodging, caused by strong winds occurs periodically in these areas. Therefore, development of dwarf plantain cultivars and elucidation of the inheritance of dwarfism is desirable. A heterozygous normal plantain cultivar (2n= 2x) with long false-internodes (19.4 ± 0.9 cm) was crossed with a homozygous wild banana (2n= 2x) with short false-internodes (6.9 ± 0.4 cm) to develop a test-cross segregating population. A total of 74 euploids (2x, 3x and 4x) were produced. Forty-one normal and 24 dwarf diploids were obtained, which fits a 2: 1 trisomic test-cross segregation ratio for one locus. Dwarfism is controlled by a single recessive gene, dw. At the tetraploid level, two normal and six dwarf hybrids were produced, suggesting that the dw locus is close to the centromere and that there is a dosage effect of the dw allele at the tetraploid level.     

Somatic chromosome elimination and meiotic chromosome pairing in the triple hybrid 6x-(Solanum acaule × S. bulbocastanum) × 2×-S. phureja

Summary Two selected hexaploid F₁ clones from the cross Solanum acaule x S. bulbocastanum were intercrossed and the resulting hybrid plants pollinated with the diploid species S. phureja, in order to obtain tetraploid triple hybrids with the same ploidy level as S. tuberosum cultivars (2n=4x=48).Apart from three trihaploids a large population of triple hybrids was obtained, showing chromosome mosaicism in root tip cells (euploid + hypoploid chromosome numbers) and a uniform, mostly hypoploid chromosome number in the pollen mother cells. It is demonstrated that somatic chromosome elimination in the early stages of development is the most probable cause.From detailed pachytene observations as well as from the chromosome associations observed at metaphase I it is evident, that there is normal pairing between the four genomes in the triple hybrids. Although S. bulbocastanum is a quite distinct species with a rigid crossability barrier with S. phureja, the chromosomes of these two species appear to have a high degree of homology. Especially the formation of quadrivalents involving all twelve groups of four homeologous chromosomes, indicated that the four parental genomes (two from S. acaule, one from S. bulbocastanum and one from S. phureja) are not differentiated to the extent of affecting normal pairing and chiasma formation. These results support the view, that the transfer of valuable characters from S. bulbocastanum to S. tuberosum cultivars is feasible even when these characters are polygenically controlled.     

Interspecific hybridization in sexual diploid Allium senescens var. minor× apomictic tetraploid A. nutans and sexual diploid A. senescens var. minor× apomictic hexaploid A. senescens

Fourteen interspecific hybrids in sexual diploid Allium senescens var. minor× apomictic tetraploid Allium nutans L. crosses, and eight interspecific hybrids in sexual diploid A. senescens var. minor× apomictic hexaploid A. senescens L. crosses were produced. The number of chromosomes was 2n= 24 in interspecific hybrids of diploid × tetraploid, and 2n= 32 in diploid × hexaploid crosses. Triploid and tetraploid interspecific hybrids showed intermediate parental morphological characteristics. Tetraploid interspecific hybrids of A. senescens var. minor×A. senescens crosses formed two groups based on leaf colour and leaf width. Seeds were formed in 11 out of 14 triploid interspecific hybrids under natural conditions. In cytological observations of parthenogenesis, three out of 12 triploid interspecific hybrids and five out of eight interspecific tetraploid hybrids were observed. Parthenogenesis ranged from 26.0% to 86.0% in five tetraploid interspecific hybrids. Non-parthenogenesis to parthenogenesis segregated in a 3:5 ratio in A. senescens var. minor×A. senescens crosses.     

Meiotic behavior in interspecific hybrids between Brachiaria ruziziensis and Brachiaria brizantha (Poaceae)

The meiotic behavior of two half-sib interspecific tetraploid (2n = 4x = 36) promising hybrids, a sexual and an apomictic one, from crosses B. ruziziensis and B. brizantha, was evaluated. Although chromosome paired predominantly as bivalents, a few tri- and quadrivalents were recorded. Results suggest that B. brizantha and B. ruziziensis are closely related and genetic recombination is expected in hybrids. Introgression of specific target genes from B. ruziziensis into B. brizantha and vice-versa may be foreseen. However, abnormalities such as irregular chromosome segregation, chromosome stickiness and abnormal cytokinesis reported in these hybrids affect pollen fertility. More than 65% of pollen grains are sterile. Since the distinctive cytological feature of these hybrids is abnormal cytokinesis, this fact suggests that both parental genomes are unable to coordinate their activities with regard to this cytological phenomenon. Deployment of such hybrids in the process of developing varieties is discussed.     

Phenotypic characterization and nuclear microsatellite analysis reveal genomic changes and rearrangements underlying androgenesis in tetraploid potatoes (Solanum tuberosum L.)

Two (di)haploids (2n = 2x = 24) and nine tetraploids (2n = 4x = 48) obtained from Solanum tuberosum through anther culture were characterized for nDNA variation, phenotypic variation and nuclear microsatellite polymorphism. Androgenic (di)haploids were also characterized for late blight resistance. The (di)haploid C-13 was derived from Indian tetraploid potato cv. Kufri Chipsona-2, while D4 from TPS (true potato seed) parental line JTH/C-107, which is an interspecific hybrid between Indian tetraploid cv. Kufri Jyoti and diploid (2n = 2x = 24) cultivated species S. phureja Juz. & Buk. IVP-35. C-13 and D4 (both male-fertile) could be distinguished from their corresponding tetraploid anther donors based on plant height, shoot number, terminal leaflet length and width, leaf ratio, anther length, pollen diameter and corolla width and radius. A complete reversal of flower color occurred in D4, and C-13 was highly resistant to late blight. Most interestingly, about 3–7% increase in nDNA content occurred in most of the anther-derived tetraploids. Both the androgenic (di)haploids and their anther donors had unique genotypes at the microsatellite loci POTM1-2, STM0015 and STM0019b. However, the nine anther-derived tetraploids shared the same allelic profiles with their anther donor JTH/C-107 at all the microsatellite loci, except at STM0019a where they were characterized by the absence of a standard donor allele (186-bp). A typical (di)haploid-specific allele was detected for the locus STWAX-2 where the standard donor alleles were replaced by a 230-bp allele in both C-13 and D4. The over-expression of microsatellite variation in D4 that also shows triallelic profiles at the microsatellite loci POTM1-2 and STM0015 can perhaps be attributed to its chimeric structure, which might have been formed through incomplete fusion of two different pro-embryos during the first steps of microspore division.