Chimeric inheritance of organelle DNA in variegated leaf seedlings from inter-subgeneric crossing of azalea

The inheritance of organelle DNA was investigated using PCR–RFLP markers in reciprocal cross combinations of inter-subgeneric azalea hybrids between evergreen azaleas (Rhododendron nakaharai and its hybrids) and fragrant deciduous azaleas (R. arborescens and R. viscosum) for the purpose of breeding fragrant evergreen azaleas. The hybrid progenies included green leaf, pale green leaf, variegated leaf and albino seedlings. Most viable green leaf seedlings had inherited ptDNA from the deciduous parent and non-viable albino and pale green leaf seedlings had inherited ptDNA from the evergreen parent. On the other hand, variegated leaf seedlings had chimerically inherited ptDNA from both parents. Their green leaf segments had ptDNA from the deciduous parent, and the pale green and white segments had biparental or maternal ptDNA depending on the progeny. In this study, we obtained interesting inter-subgeneric azalea hybrid progenies that had chimerically inherited organelle DNA and had different colored leaf segments corresponding to the composition of ptDNA from each parent. These results suggest that variegated leaf progenies with chimeric ptDNA from both parents can be subsistent, whereas albino seedlings resulting from plastome–genome incompatibility between the plastid genome from evergreen azalea and the nuclear genome from deciduous azalea are non-viable.     

Development of multiplex PCR markers to distinguish evergreen and deciduous azaleas

We successfully developed new PCR‐based markers for analysing the DNA inheritance patterns of vigorous inter‐subgeneric hybrids of azalea. The hybrids were between evergreen azaleas (Rhododendron nakaharae and its hybrids) and fragrant deciduous azaleas (Rhododendron arborescens and Rhododendron viscosum). We determined the sequence of the nuclear internal transcribed spacer (ITS) region in evergreen and deciduous azaleas, and found a deletion of five base pairs (bp) in evergreen azaleas compared with deciduous azaleas. Sequences for the chloroplast matK‐trnK region were obtained from GenBank. Primers for the multiplex PCRs were designed using these sequence data. In the multiplex PCR of the ITS region, evergreen azaleas showed a specific band of 140 bp and deciduous azaleas showed a specific band of 800 bp. In the multiplex PCR of the matK‐trnK region, evergreen azaleas showed a specific band of 560 bp and deciduous azaleas showed a specific band of 300 bp. These results indicate that multiplex PCR methods are useful to distinguish evergreen and deciduous azaleas easily and rapidly.     

Effectiveness of three-way crossing for the breeding of yellow-flowered evergreen azalea

For the purpose of obtaining yellow-flowered evergreen azaleas, single and three-way interspecific crosses between evergreen azaleas and Rhododendron japonicum f. flavum were compared. In single cross, although many seeds were obtained from R. eriocarpum × R. japonicum f. flavum, the seedlings from immature ovule were albino or pale-green plants. They were withered away at acclimation. In R. kiusianum × R. japonicum f. flavum, green seedlings were observed, but number of ovules contained in one capsule was very small. In three-way cross, (R. kiusianum × R. eriocarpum) × R. japonicum f. flavum, many viable seeds were contained in one capsule, and about 15% of the seedlings were green leaf colored. These results suggest that three-way cross is one of the useful breeding methods for creating the new yellow-flowered evergreen azalea. Isozyme analysis revealed that most of seedlings were zygotic origin without distinction of their leaf colors, but some green seedlings indicated the same genotypes as the seed parents. Therefore, it is important to confirm hybridity of the seedlings by isozyme technique. This revised version was published online in August 2006 with corrections to the Cover Date.     

Resynthesis of rapeseed (Brassica napus L.): A comparison of sexual versus somatic hybridization

Sexual and somatic Brassica napus hybrids produced from the same parental plants were compared. Sexual crosses between a white-flowered, self-compatible broccoli selection (B. oleracea var. italica, cc genome) as the maternal parent and a flowering pak choi accession (B. chinensis, aa genome) yielded one unique spontaneous hybrid and four hybrids through embryo rescue. Thirty-nine somatic hybrids were recovered from a protoplast fusion experiment. Hybridity was confirmed by morphology, isozyme expression, flow cytometry, and DNA hybridization. Sexual and somatic hybrids exhibited differences in leaf morphology, flower colour, flowering habit, and organellar inheritance. Sexual hybrids were all fertile amphidiploids (2n = 38, aacc) following spontaneous chromosome doubling. All somatic hybrids had high nuclear DNA contents; most were probably hexaploids (aaaacc or aacccc) from the fusion of three portoplasts. Two initially sterile hexaploid (aaaacc) regenerates eventually set selfed seed after the loss of the putative extra aa genome following regrowth from axillary buds. A bias toward inheritance of B. chinensis chloroplasts was observed with somatic hybrids.     

Intergeneric symmetric and asymmetric somatic hybridization in Festuca and Lolium

Summary Intergeneric symmetric and asymmetric somatic hybrids have been obtained by fusion of metabolically inactivated protoplasts from embryogenic suspension cultures of tall fescue (Festuca arundinacea Schreb.) and unirradiated or 10–500 Gy-irradiated protoplasts from non-morphogenic cell suspensions of Italian ryegrass (Lolium multiflorum Lam.). Genotypically and phenotypically different somatic hybrid Festulolium mature flowering plants were regenerated.Species-specific sequences from F. arundinacea and L. multiflorum being dispersed and evenly-represented in the corresponding genomes were isolated and used for the molecular characterization of the nuclear make-up of the intergeneric, somatic Festulolium plants recovered. The irradiation of Italian ryegrass protoplasts with 250 Gy X-rays prior to fusogenic treatment favoured the unidirectional elimination of most or part of the donor chromosomes. Irradiation of L. multiflorum protoplasts with 500 Gy produced highly asymmetric (over 80% donor genome elimination) nuclear hybrids and clones showing a complete loss of donor chromosomes.The RFLP analysis of the organellar composition in symmetric and asymmetric tall fescue (+) Italian ryegrass regenerants confirmed their somatic hybrid character and revealed a bias towards recipient-type organelles when extensive donor nuclear genome elimination had occurred.Approaches aimed at improving persistence of ryegrasses based on asymmetric somatic hybridization with largely sexually-incompatible grass species (F. rubra and Alopecurus pratensis), and at transferring the cytoplasmic male sterility trait by intra- and inter-specific hybridization in L. multiflorum and L. perenne, have been undertaken.Abbreviations cpDNA chloroplast DNA - CMS cytoplasmic male sterility - 2,4-D 2,4-dichlorophenoxy-acetic acid - IOA iodoacetamide - mtDNA mitochondrial DNA - RFLP restriction fragment length polymorphism     

Adaptation to iron deficiency and high pH in evergreen azaleas (<Emphasis Type="Italic">Rhododendron</Emphasis> spp.): potential resources for breeding

The growth of evergreen azaleas (Rhododendron spp.) can be altered by iron (Fe) chlorosis when plants are cultivated in a neutral-alkaline substrate. In this study, morphological and physiological responses to alkalinity and Fe deficiency were evaluated in five diploid Japanese azaleas to assess their potential as resources for breeding. R. obtusum ‘Kirin’, R. indicum ‘Shinsen’, R. × pulchrum ‘Sen-e-oomurasaki’, R. indicum ‘Osakazuki’, and R. ripense were pot cultivated in a peat-based substrate for 10 weeks, in acid and alkaline growing media with both adequate and inadequate Fe nutrition. Plant performance was generally affected by high pH of the substrate, while Fe deficiency by itself influenced few of the evaluated parameters, possibly due to the complex adaptive response mechanisms of these slow growing ornamental shrubs. According to the biochemical and physiological variations recorded on a long period of cultivation, R. indicum ‘Osakazuki’ reported the best performance. This azalea could be a valuable resource for breeders.     

Production and genetic evaluation of interspecific hybrids within the genus Sambucus

Attempts were made to produce hybrids among various Sambucus species in order to change the plant structure of S. nigra, a medicinal plant of increasing agricultural importance. The first successful genetic recombination, involving four species, was conducted after the introduction of S. javanica from a South Pacific island. A series of genetic analyses of the involved species and some relatives was performed in order to obtain reliable data on genetic similarity among parental species and to confirm the hybrid nature of putative hybrids. The nuclear 2C DNA content revealed by flow cytometry of nine species showed limited variation, the greatest being in S. nigra (28.6 pg) and the smallest in S. caerulea (23.9 pg). Maximum parsimony analysis of seven Sambucus species was used to form two dendrograms, one based on the sequenced nuclear ribosomal DNA (nrDNA) internal transcribed spacer (ITS) region and the other based on the cpDNA trnT‐trnF region. The results showed similar clustering into distinct groups; on both dendrograms S. nigra and S. javanica ES were closely related. Analysis of hybrid origin was based on the sequenced cpDNA region, genome size differences and PCR–RFLP analysis of the nrDNA ITS region. The seed parent and maternal origin of cpDNA was confirmed for all hybrids, while the pollen parent was confirmed in 21 of 22 putative hybrid plants tested.     

Genetic analysis of <Emphasis Type="Italic">Jatropha</Emphasis> species and interspecific hybrids of <Emphasis Type="Italic">Jatropha curcas</Emphasis> using nuclear and organelle specific markers

The present study aims at characterization of Jatropha species occurring in India using nuclear and organelle specific primers for supporting interspecific gene transfer. DNA from 34 accessions comprising eight agronomically important species (Jatropha curcas, J. gossypifolia, J. glandulifera, J. integerrima, J. podagrica, J. multifida, J. villosa, J. villosa. var. ramnadensis, J. maheshwarii) and a natural hybrid, J. tanjorensis were subjected to molecular analysis using 200 RAPD, 100 ISSR and 50 organelle specific microsatellite primers from other angiosperms. The nuclear marker systems revealed high interspecific genetic variation (98.5% polymorphism) corroborating with the morphological differentiation of the species used in the study. Ten organelle specific microsatellite primers resulted in single, discrete bands of which three were functional disclosing polymorphism among Jatropha species. The PCR products obtained with organelle specific primers were subjected to sequence analysis. PCR products from two consensus chloroplast microsatellite primer pairs (ccmp6 and 10) revealed variable number of T and A residues in the intergenic regions of ORF 77–ORF 82 and rp12–rps19 regions, respectively in Jatropha. Artificial hybrids were produced between J. curcas and all Jatropha species used in the study with the exception of J. podagrica. Characterization of F₁ hybrids using polymorphic primers specific to the respective parental species confirmed the hybridity of the interspecific hybrids. Characterization of both natural and artificially produced hybrids using chloroplast specific markers revealed maternal inheritance of the markers. While the RAPD and ISSR markers confirmed J. tanjorensis as a natural hybrid between J. gossypifolia and J. curcas, the ccmp primers (ccmp6 and 10) unequivocally established J. gossypifolia as the maternal parent. Evaluation of backcross interspecific derivatives of cross involving J. curcas and J. integerrima indicate scope for prebreeding and genetic enhancement of Jatropha curcas through interspecific hybridization.     

Synthesis of somatic hybrids (RCBB) by fusing heat-tolerant Raphanus sativus (RR) and Brassica oleracea (CC) with Brassica nigra (BB)

Brassica carinata (BBCC), a potential oilseed crop for dry land agriculture, is sensitive to high temperatures during germination and early stages of growth, which thereby restricts the possibility of using the residual soil moisture available after the rainy season for its cultivation. To overcome this problem, a three‐genome hybrid, RCBB, was synthesized using Raphanus sativus (RR) and Brassica oleracea (CC) as donor sources for the desired heat tolerance. Protoplasts of RC hybrids obtained through sexual crosses between R. sativus (female) and B. oleracea (male) were fused with protoplasts of Brassica nigra (BB) to produce RCBB somatic hybrids. The hybrid colonies regenerated with an average frequency of 7.6%. Twelve out of 36 hybrids grown to maturity were characterized for their nuclear and organelle genomes. While all the hybrids showed the presence of B. nigra chloroplasts, 10 of the hybrids showed B. nigra‐specific mitochondria and two had Raphanus‐spedfic mitochondria. The somatic hybrids could be backcrossed to B. carinata.     

Cytogenetical and molecular investigations on somatic hybrids of Sinapis alba and Brassica juncea and their backcross progeny

Somatic hybrids of Sinapis alba+Brassica juncea (Sal Sal AABB) were synthesized by protoplast electrofusion. They were true genomic allopolyploids since they possessed 60 chromosomes, i.e. the sum of S. alba (2n= 24) and B. juncea (2n= 36) chromosomes. Chromosome pairing was predominantly bivalent with the occasional occurrence of multivalents in the pollen mother cells at diakinesis and metaphase I. Hybrids were completely pollen-sterile, but produced seeds on back-crossing with B. juncea and B. campestris. A total of 37 BC₁ plants were raised from two somatic hybrids (JS-1 and JS-2) and 24 of these were analysed cytologically. The 22 plants originating from the pollinations of somatic hybrids with B. juncea showed a chromosome configuration of 18II+12I and had 42–86% pollen fertility. Two plants from the backcrosses of the somatic hybrid with B. campestris formed 10II +20I, and had 0–4% fertile pollen. Total DNA analysis by probing with pTA71 carrying a full-length 18S–25S rDNA fragment of the wheat nuclear genome revealed that the two somatic hybrids possessed all the characteristic bands of both the species, confirming their hybridity. Probing with the mitochondrial coxI and atp9 genes indicated mitochondrial genome recombination in the hybrids. Hybridization with chloroplast-specific psbD indicated that both the somatic hybrids possessed the cp genome of S. alba origin.     

Stomatal Density and Length for Breeding of Evergreen Azaleas (Rhododendron simsii Planch.)

The hentability of stomatal density and stomatal length has been studied for four evergreen azalea (Rhododendron simsii Planch.) cultivars with the aim to evaluate the possibilities of stomatal characteristics of parental plants for increasing the crown diameter – hence yield – of the offspring., A nearly complete diallel cross with reciprocals has been carried out with the four parental cultivars. Highly significant general combining ability (GCA) values of the parents were found for stomatal length and density, but not for the crown diameter. GCA values of crown diameters did not correspond with GCA values of either stomatal length or stomatal density. Therefore it was concluded that utilizing stomatal characteristics as a selection tool for an increased crown diameter (yield) in azalea., may not be feasible. Differences in crown diameter could not be explained from correlations with either stomatal density or stomatal length.     

Production of Eruca-Brassica Hybrids by Embryo Rescue

Allied species of crop Brassicas have potential value as donors of useful nuclear/organelle genomes. Eruca sativa, a member of subtribe Brassicineae, is resistant to white rust and well adapted to drought. Attempts to hybridize it with Brassica campestris by conventional methods were unsuccessful. However, hybrids were obtained by embryo rescue and the hybrid embryos were found to produce numerous secondary embryos in upto 7 to 8 subcultures. Plantlets developed from them exhibited morphological characteristics of both parents. The chromosome number of 2n = 42 showed that they were amphidiploids. The plants were allotctraploids (2n = 42) and showed 21 bivalents at M₁ of meiosis. Hybridization of total DNA of the hybrids with two probes, a B. campestris tandem repeat DNA and 18s ribosomal DNA of wheat showed that it was derived from the genome of both parents. The hybrids are self-fertile and show the same high fertility even in A³ generation. They were selfed or crossed with B. juncea, B. campestris and B. nigra to screen for useful agronomical traits. Six more embryos of E. saliva × B. campestris have been obtained and their growth is being studied. Preliminary small scale field trials indicate that the selfed hybrid is comparable in yield with a high yielding commercial variety of B. juncea.     

Transfer of disease resistance within the genus Brassica through asymmetric somatic hybridization

Summary Asymmetric somatic hybrid plants between Brassica napus L. (oilseed rape genome AACC) and a transgenic line of Brassica nigra L. Koch (black mustard genome BB) were tested for their resistance against rapeseed pathogens Phoma lingam (black leg disease) and Plasmodiophora brassicae (club root disease). The transgenic B. nigra line used (hygromycin-resistant, donor) is highly resistant to both fungi, whereas B. napus (recipient) is highly susceptible. The asymmetric somatic hybrids were produced using the donor-recipient fusion method (with X-irradiation of donor protoplasts) reported by Zelcer et al. (1978) for the production of cybrids. Using hygromycin-B for selection, a total of 332 hybrid calli were obtained. Regenerants, resistant or susceptible to both diseases, were selected. Many hybrids expressed resistance to only one pathogen. Dot blot experiments showed that the asymmetric hybrid plants contained varying amounts of the donor genomic DNA. Furthermore, a correlation was detected between the radiation dose and the degree of donor DNA elimination.     

A stable cytoplasmic male-sterile line of Brassica juncea carrying restructured organelle genomes from the somatic hybrid Trachystoma ballii+B. juncea

A cytoplasmic male-sterile (CMS) line of Brassica juncea has been developed by combining the cytoplasm originating from the somatic hybrid Trachystoma ballii+B. juncea, and the nucleus of B. juncea cv. Pusa Bold by repeated backcrossing. Male-sterile plants closely resembled the normal fertile B. juncea in general morphology, but had delayed flowering (5–7 days) when compared with fertile ‘Pusa Bold’ which flowered in 45 days. Stamens of the male-sterile line were transformed into petaloid structures. Pollen abortion occurred after tetrad formation. Female fertility of the male-sterile line was normal. Molecular analysis of organelle genomes indicated extensive mitochondrial DNA recombinations in the CMS line. Preliminary analysis of the chloroplast genome of the CMS line also indicated chloroplast DNA recombination.     

Intergeneric crosses for the transfer of resistance to the beet cyst nematode from Raphanus sativus to Brassica napus

Summary The possibilities to transfer important traits and in particular resistance to the beet cyst nematode (Heterodera schachtii, abbrev. BCN) from Raphanus sativus to Brassica napus were investigated. For these studies B. napus, R. sativus, the bridging hybrid ×Brassicoraphanus (Raparadish) as well as offspring of the cross ×Brassicoraphanus (Raparadish) ×B. napus were used. Reciprocal crosses between B. napus and R. sativus were unsuccessful, also with the use of embryo rescue. Crosses between ×Brassicoraphanus as female parent and B. napus resulted in a large number of F₁ hybrids, whereas the reciprocal cross yielded mainly matromorphic plants. BC₁, BC₂ and BC₃ plants were obtained from backcrosses with B. napus, which was used as the male parent. F₁ hybrids and BC plants showed a large variation for morphology and male and female fertility. Cuttings of some F₁ and BC₁ plants, obtained from crosses involving resistant plants of ×Brassicoraphanus, were found to possess a level of resistance similar to that of the resistant parent. These results and indications for meiotic pairing between chromosomes of genome R with those of the genomes A and/or C suggest that introgression of the BCN-resistance of Raphanus into B. napus may be achieved.     

<Emphasis Type="Italic">Nicotiana debneyi</Emphasis> has a single dominant gene causing hybrid lethality in crosses with <Emphasis Type="Italic">N. tabacum</Emphasis>

To elucidate the genetic mechanism of hybrid lethality observed in hybrids between cultivated tobacco, Nicotiana tabacum, and wild tobacco species in the section Suaveolentes, genetic analyses were conducted through the triple cross of the hybrids of wild species, including N. benthamiana and N. fragrans, and N. tabacum. N. benthamiana and N. fragrans produced only viable hybrids after crossing with N. tabacum. Subsequently, N. benthamiana and N. fragrans were crossed with N. africana, N. debneyi, and/or N. suaveolens, which produced inviable hybrids after crossing with N. tabacum. Hybrids of the wild species were obtained from four of the six cross combinations. Only when hybrid plants of N. debneyi × N. fragrans, whose hybridity was confirmed by morphological characteristics, random amplified polymorphic DNA analysis, and chromosome observation, were crossed with N. tabacum, triple hybrids were obtained and segregated 1:1 (lethal:viable). Based on these results, a single dominant gene, designated Hybrid Lethality A1 (HLA1), in N. debneyi was found to control hybrid lethality by the interaction with gene(s) on the Q chromosome in N. tabacum. This represents the first report to identify a causal gene for hybrid lethality in the genus Nicotiana.     

Induction of novel organelle DNA variation and transfer of resistance to frost and Verticillium wilt in Solanum tuberosum through somatic hybridization with 1EBN S. commersonii

Somatic hybridization can be used to induce genetic variability in plastidial and mitochondrial genomes, and transfer multiple uncloned genes across sexual barriers. Somatic hybrids were produced between a dihaploid clone of the common potato, S. tuberosum subsp. tuberosum, and the wild sexually incongruent diploid species S. commersonii. Fusion products were selected on the basis of callus growth and regeneration in vitro. Genome composition of putative somatic hybrids was determined by flow cytometric analysis of nuclear DNA content, RAPD analysis, and Southern analysis with probes specific to organellar DNA. All regenerated fusion products proved to be hybrids based on RAPD analysis. Seventy per cent of somatic hybrids were (near) tetraploids, 22% (near) hexaploids and 8%(near) octoploids. A high correlation was found between the nuclear DNA content determined by flow cytometry and the number of chloroplasts in stomata guard cell pairs. Somatic hybrids inherited the parental plastids in a random manner. On the contrary, they preferentially inherited the mitochondrial DNA fragments of S. tuberosum. The majority of them had a rearranged mitochondrial genome with fragments from both parents. Hybrids were highly vigorous and morphologically more similar to the cultivated than to the wild parent, produced tubers on long stolons under long photoperiod conditions, showed a high degree of flowering, but did not produce pollen. In addition, somatic hybrids were generally more resistant to frost and Verticillium wilt than the cultivated parent, indicating the introgression of relevant resistance genes from the wild species into the genetic background of S. tuberosum. This revised version was published online in July 2006 with corrections to the Cover Date.     

Production and characterization of intergeneric somatic hybrids between Moricandia arvensis and Brassica oleracea

Somatic hybrids were produced between Moricandia arvensis (MaMa, 2n= 28) and Brassica oleracea (CC, 2n= 18) through cell fusion and then characterized by analysing their morphology, cytology, DNA constitution, leaf anatomy and seed fertility. Cell fusion was carried out between greenish protoplasts isolated from the mesophyll of M. arvensis and colourless ones from hypocotyls of B. oleracea. Three plants were generated from one shoot via cuttings and acclimatized in vivo. They closely resembled each other in morphology, exhibiting traits intermediate between the parental species. They were confirmed to be amphidiploids by mitotic and meiotic analyses, being 2n= 46 (MaMaCC), with pollen fertility of about 50%, which was enough to develop the subsequent progenies. Anatomical analysis of the for leaf tissue showed that the bundle sheath cells of the somatic hybrids contained some centripetally arranged organelles, like those of M. arvensis. The hybridity was also confirmed by random amplified polymorphic DNA analysis. Both chloroplast DNA and mitochondrial DNA of the somatic hybrids were estimated to be derived from M. arvensis. In leaf anatomy, the somatic hybrid showed the C₃‐C₄ intermediate trait as in M. arvensis. Many progenies resulted from backcrossing with parental species. The somatic hybrids are expected to be used as bridging plant material to introduce the C₃‐C₄ intermediate trait into Brassica crop species.     

Interspecific hybridization between Cyclamen persicum Mill, and C. purpurascens Mill.

In order to introduce valuable traits from wild species of Cyclamen into C. persicum cultivars, crosses were made between C. persicum‘Reinweiß’and C. purpurascens. Crossing barriers between C. persicum‘Reinweiß’and C. purpurascens were due to late-acting incompatibility reactions. Interspecific hybrids were obtained by using ovary culture. The highest number of embryos was achieved from placentas excised 21 days and 35 days after pollination and transferred to Murashige-Skoog (M.S.)–medium containing 6% sucrose and 1% agar. The hybrids showed a habit and a chromosome number intermediate between the parents. The fragrant flowers were pale red-purple. The chromosome number in root tips was determined as 2n= 41 while in C. persicum it is 2n= 48 and in C. purpurascens 2n= 34. Due to the different parental chromosome sizes, chromosomes of distinct size were still observed in the hybrid. Pollen viability varied between 0.3 and 34.0%. Parents and interspecific hybrids also showed differences in the DNA content of leaf tissue. Flow cytometric analyses were useful in the early identification of hybrids.     

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