Regeneration of somatic hybrid plants formed between Lycopersicon esculentum and Solatium rickii

Summary A single somatic hybrid callus clone was identified following the fusion of Lycopersicon esculentum protoplasts and Solanum rickii suspension culture protoplasts. The hybrid nature of the callus and the plants regenerating from it was determined by assaying phosphoglucomutase-2 isozyme expression. The chloroplast genome present in four somatic hybrid plants was characterized by probing digests of total DNA with nick translated L. esculentum chloroplast DNA(cpDNA). All four somatic hybrid plants had inherited S. rickii cpDNA. Two clones of plant mitochondrial DNA (mtDNA), soybean 18S and 5S rDNA and maize cytochrome oxidase subunit II were used to characterize the mtDNA present in total DNA digests of four somatic hybrid plants. In both cases, the somatic hybrid plants had inherited most but not all of the S. rickii specific fragments, but none of the L. esculentum specific fragments.     

Regeneration of somatic hybrid plants formed between Lycopersicon esculentum and L. pennellii

Summary Somatic hybrid plants have been regenerated following polyethylene glycol mediated fusion of leaf mesophyll protoplasts from tomato and protoplasts from Lycopersicon pennellii callus. Three different cultivars of tomato were used as sources of protoplasts: Early Girl, Manapal, and UC82B. Fusions were performed between protoplasts of these tomato cultivars and protoplasts of L. pennellii, and between protoplasts of the cultivars and protoplasts of L. pennellii that had been exposed to 3 or 6 krads of gamma radiation. Somatic hybrid plants were identified on the basis of heterozygous isozyme banding patterns, and leaf and flower morphology. Somatic hybrid plants were regenerated following fusion of tomato protoplasts with either untreated or irradiated L. pennellii protoplasts. All were heterozygous for isozyme loci on five different chromosomes. Regenerated somatic hybrids showed inheritance of either or both parental chloroplast genomes, but predominantly the L. pennellii mitochondrial genome. The regenerated somatic hybrid plants exhibited reduced fertility, less than 20% viable pollen. A total of 34 somatic hybrid calli were identified. Of these, 21 regenerated shoots, and 7 produced seed following manual pollinations.     

Examination of genome stability in cultured Lycopersicon

The genetic stability of plants regenerated from either mesophyll protoplasts or leaf slices of the F1 hybrid between Lycopersicon esculentum and L. pennellii was assayed by comparing the ploidy level, leaf morphology and isozyme patterns of the regenerants with their somatic parents. Regenerants from protoplasts were predominantly tetraploid, regenerants from leaf slices were predominantly diploid; both classes of regenerants had isozyme patterns identical to those of the parent plant. Callus was analyzed that grew up from cultures containing fused protoplasts from either irradiated or untreated protoplasts of L. esculentum and L. pennellii. The L. pennellii cell line used was 18 months old and could no longer regenerate. Out of 75 calli scored at 3 isozyme loci, 51 were heterozygous at only one or two of the loci. Irradiation of the two parental lines was not necessary to produce fusion products exhibiting asymmetric expression of parental genes.Abbreviations Got-2 glutamate oxaloacetate transaminase-2 - Pgi-1 phosphoglucoisomerase-1 - Pgm-2 phosphoglucomutase-2     

Asymmetric somatic hybrid plants between an interspecific Lycopersicon hybrid and Solanum melongena

Summary Asymmetric somatic hybrid plants were obtained by a modified PEG/DMSO fusion procedure between protoplasts derived from suspension cells of an interspecific tomato hybrid, Lycopersicon esculentum x L. pennellii, and mesophyll protoplasts of Solanum melongena, eggplant. The tomato hybrid was previously transformed with Agrobacterium tumefaciens and contained the kanamycin-resistance marker gene. Prior to fusion, the donor protoplasts of the tomato hybrid were gamma irradiated at 9.0 krad. Thus, non-division of irradiated tomato hybrid protoplasts coupled with kanamycin sensitivity of eggplant enabled selection of somatic cell hybrids. Forty-nine calli selected post-fusion regenerated leaf-like structures in the presence of 50 mg/l kanamycin. However, only four of the 49 calli regenerated intact shoots which rooted in the presence of 50 mg/l kanamycin and were later transferred to the greenhouse. Analysis of phosphoglucoisomerase and peroxidase isozymes, and Southern hybridization with a nuclear-specific pea 45 S ribosomal RNA gene confirmed somatic hybrid status. Cytology revealed that the four hybrid plants had chromosome numbers of 45, 60, 42 and 57, respectively; they were all sterile.     

The role of irradiation dose and DNA content of somatic hybrid calli in producing asymmetric plants between an interspecific tomato hybrid and eggplant

Highly asymmetric somatic hybrid plants were obtained by PEG/DMSO fusion of gamma-irradiated mesophyll protoplasts of the kanamycin-resistant (KmR⁺) interspecific hybrid Lycopersicon esculentum x L. pennellii (EP) with mesophyll protoplasts of Solanum melongena (eggplant, E). Elimination of the EP chromosomes was obtained by irradiating the donor genome with different doses of gamma rays (100, 250, 500, 750 and 1000 Gy). The selection of somatic hybrid calli was based on kanamycin resistance; EP and E protoplasts did not divide due to the irradiation treatment and sensitivity to kanamycin, respectively. KmR⁺ calli were recovered following all irradiation doses of donor EP protoplasts. The hybrid nature of the recovered calli was confirmed by PCR amplification of the NptII gene, RAPD patterns and Southern hybridizations using potato ribosomal DNA and pTHG2 probes. Ploidy levels of calli confirmed as hybrid were further analyzed by flow cytometry. Such analyses revealed that the vast majority of hybrid calli that did not regenerate shoots were 5–9n polyploids. The three asymmetric somatic hybrid plants obtained were regenerated only from callus with a ploidy level close to 4n, and such calli occurred only when the donor EP had been exposed to 100 Gy. The amount of DNA in somatic hybrid calli, from 100-Gy exposure, was found by dot blot hybridization with the species-specific probe, pTHG2, to be equivalent with 3.1–25.8% of the tomato genome. Thus, DNA contained in 3.8–13.2 average-size tomato chromosomes was present in these hybrid calli. The asymmetric somatic hybrid plants had the eggplant morphology and were regenerated from one hybrid callus that contained an amount of tomato DNA equivalent to 6.29 average-size tomato chromosomes.     

Somatic hybrid plants between the forage legumes Medicago sativa L. and Medicago arborea L.

Interspecific somatic hybrid plants were obtained by symmetrical electrofusion of mesophyll protoplasts of Medicago sativa with callus protoplasts of Medicago arborea. Somatic hybrid calli were picked manually from semi-solid culture medium after they were identified by their dual color in fluorescent light. Twelve putative hybrid calli were selected and one of them regenerated plants. The morphogenesis of the somatic hybrid calli was induced by the synthetic growth regulator 1,2 benzisoxazole-3-acetic acid. Somatic hybrid plants showed intensive genome rearrangements, as evidenced by isozyme and RFLP analysis. The morphology of somatic hybrid plants was in general intermediate between the parents. The production of hybrids by protoplast fusion between sexually incompatible Medicago species is related to the in vitro respon siveness of the parental protoplasts. The possibility of using somatic hybrid plants in alfalfa breeding is discussed.     

Molecular analysis of the nuclear organellar genotype of somatic hybrid plants between tomato (Lycopersicon esculentum) and Lycopersicon chilense

Summary Somatic hybrid plants were recovered following fusion of leaf mesophyll protoplasts isolated from tomato (Lycopersicon esculentum) cultivar UC82 with protoplasts isolated from suspension cultured cells of L. chilense, LA 1959. Iodoacetate was used to select against the growth of unfused tomato protoplasts. Two somatic hybrids were recovered in a population of 16 regenerants. No tomato regenerants were recovered; all of the non-hybrid regenerants were L. chilense. The L. chilense protoplast regenerants were tetraploid. The hybrid nature of the plants was verified using species-specific restriction fragment length polymorphisms for the nuclear, chloroplast and mitochondrial genomes. The somatic hybrids had inherited the chloroplast DNA of the tomato parent, and portions of the mitochondrial DNA of the L. chilense parent. The somatic hybrids formed flowers and developed seedless fruit.     

Non-random inheritance of organellar genomes in symmetric and asymmetric somatic hybrids between Lycopersicon esculentum and L. pennellii

Summary The organization of the mitochondrial genome and the genotype of the chloroplast genome was characterized using restriction fragment length polymorphisms in a population (82 individuals) of symmetric and asymmetric somatic hybrids of tomato. The protoplast fusion products were regenerated following the fusion of leaf mesophyll protoplasts of Lycopersicon esculentum (tomato cv UC82) with suspension cell protoplasts of L. pennellii that had been irradiated with 5, 10, 15, 25, 50, or 100 kRads from a gamma source. The chloroplast genome in the somatic hybrids showed a random pattern of inheritance, i.e., either parental genome was present in equal numbers of regenerants, while in asymmetric somatic hybrids, the chloroplast genotype reflected the predominant nuclear genotype, i.e., tomato. The mitochondrial genome in the symmetric somatic hybrids showed a non-random pattern of inheritance, i.e., predominantly from the L. pennellii parent; asymmetric somatic hybrids had more tomato-specific mitochondrial sequences than symmetric somatic hybrids. The non-random inheritance of the chloroplast and mitochondrial DNA in these tomato protoplast fusion products appears to be influenced by the nuclear background of the regenerant.     

Somatic hybridization between selected Lycopersicon and Solanum species

Summary Mesophyll protoplasts of an interspecific Lycopersicon esculentum Mill, (tomato) x Lycopersicon pennellii hybrid plant (EP) were fused with callus-derived protoplasts of Solanum lycopersicoides Dun. using a modified PEG/DMSO procedure. The EP plant was previously transformed by Agrobacterium tumefaciens which carried the NPTII and nopaline synthase genes. Protoplasts were plated at 10⁵/ml in modified KM medium and 16 days post-fusion 25 ug/ml kanamycin was added to the culture medium. During shoot regeneration, 212 morphologically similar putative somatic hybrids were delineated visually from kanamycin resistant EP's. Forty-eight shoots, randomly selected among the 212, were further verified as somatic hybrids by their leaf phosphoglucoisomerase heterodimer isozyme pattern. However, the resulting plants were virtually pollen sterile. In a second fusion, mesophyll protoplasts of Solanum melongena (eggplant) were fused with EP callus-derived protoplasts. Using the same fusion and culture procedure, only two dark green calli were visually selected among the pale green parental EP and verified as somatic cell hybrids by several isozyme patterns. These two calli have produced only leaf primordia in one and half years on regeneration medium.Abbreviations ABA abscisic acid - BAP 6 benzylaminopurine - 2,4-D 2,4 dichlorophenoxy acetic acid - DMSO dimethyl sulfoxide - GA₃ gibberellic acid - GOT glutamate oxaloacetate - IAA indoleacetic acid - IBA indolebutyric acid - IDH isocitrate dehydrogenase - MDH malate dehydrogenase - MES morpholinoethane-sulfonic acid - PEG polyethylene glycol - 6-PGDH 6 phosphogluconate dehydrogenase - PGI phosphoglucoisomerase     

Donor chromosome elimination and organelle composition of asymmetric somatic hybrid plants between an interspecific tomato hybrid and eggplant

Morphology, the extent of elimination of donor chromosomes and the organelle composition of highly asymmetric somatic hybrid plants between a interspecific tomato hybrid Lycopersicon esculentum x L. pennellii (EP) as donor and a Solarium melongena, eggplant (E), recipient, were studied. Morphologically, the somatic hybrids most resemble eggplant but, due to polyploidy, growth is slower relative to both fusion parents. The somatic hybrids produce flowers that are characterized by abnormal styles, stigmas and by anthers which do not produce pollen. Limited amounts of donor EP genomic DNA were found in the three somatic hybrid plants (H18-1, H18-2 and H18-3), by dot-blot hybridization with probe pTHG2, equivalent to 6.23,5.41, and 5.95% EP, respectively. These percentages translated to the presence of 3.59, 2.90 and 3.19 average-size EP chromosomes in plants H1 8-1,-2 and-3, respectively. RFLP determination of L. esculentum- and L. pennellii-specific chromosomes revealed that only fragments of eight to ten out of the 24 EP chromosomes (EP has 12 L. esculentum and 12 L. pennellii chromosomes) are present in the asymmetric somatic hybrid plants. Loci of L. esculentum and L. pennellii were evenly represented in plants H18-1, -2, and -3: four to five from L. esculentum and four to five from L. pennellii. All somatic hybrid plants retained locus TG22, chromosome 4, from both EP species. Although the regeneration of plants, H18-1, -2 and-3 was from one callus, loci TG31 and TG79 of L. esculentum chromosome 2 and L. pennellii chromosome 9, respectively, were missing in hybrid plant H18-1. The three somatic hybrid plants all had chloroplast DNA fragments specific for S. melongena. The mitochondrial genome (mtDNA) in the asymmetric somatic hybrids showed predominantly the pattern of eggplant; however, some eggplant-specific polymorphic bands were not present in the three plants.     

Production of a highly asymmetric somatic hybrid between rice and Zizania latifolia (Griseb): evidence for inter-genomic exchange

 A highly asymmetric and fertile somatic hybrid plant was obtained via protoplast fusion in an intergenric combination. Gamma-ray-irradiated Zizania latifolia (Griseb). Turcz. ex Stapf mesophyll protoplasts were electrofused with idoacetamide-inactivated rice protoplasts derived from a 2-month-old suspension cell culture. Two of the six putative hybrid calli regenerated plants. Cytological observation showed that the somatic chromosome numbers of both plants were the same as the rice parent (2n=24). Nevertheless, the hybrid nature and inter-genomic exchange events of one of the plants, i.e. SH6 (SH for somatic hybrid), were confirmed by Southern analysis using both total genomic DNA and moderate-copy, Z. latifolia-abundant DNA sequences as probes; in both cases, parental specific and/or new intergenomic recombinant hybridization fragments were detected. In both plant and seed morphology, the hybrid (SH6) was distinct from its rice parental cultivar, as well as from the wild donor species, Z. latifolia. Received: 15 August 1998 / Accepted: 30 September 1998     

Rapid chloroplast segregation and recombination of mitochondrial DNA in Brassica cybrids

Summary Brassica cybrids were obtained after fusing protoplasts of fertile and cytoplasmic male sterile (CMS) B. napus lines carrying the original b. napus, and the Ogura Raphanus sativus cytoplasms, respectively. Iodoacetate treatment of the fertile line and X-irradiation of the CMS line prevented colony formation from the parental protoplasts. Colony formation, however, was obtained after protoplast fusion. Hybrid cytoplasm formation was studied in 0.5 g to 5.0 g calli grown from a fused protoplast after an estimated 19 to 22 cell divisions. Chloroplasts and mitochondria were identified in the calli by hybridizing appropriate DNA probes to total cellular DNA. Out of the 42 clones studied 37 were confirmed as cybrids. Chloroplast segregation was complete at the time of the study. Chloroplasts in all of the cybrid clones were found to derive from the fertile parent. Mitochondrial DNA (mtDNA) segregation was complete in some but not all of the clones. In the cybrids, mtDNA was different from the parental plants. Physical mapping revealed recombination in a region which is not normally involved in the formation of subgenomic mtDNA circles. The role of treatments used to facilitate the recovery of cybrids, and of organelle compatibility in hybrid cytoplasm formation is discussed.     

Molecular analysis of the extent of asymmetry in two asymmetric somatic hybrids of tomato

Summary Two somatic hybrid plants generated from a single fusion event between Lycopersicon esculentum and irradiated L. pennellii protoplasts have been analyzed at the molecular level. Over 30 loci have been analyzed using isozymes and RFLPs. All loci tested on chromosomes 2–10 were heterozygous, while those loci on chromosome 12 were homozygous L. pennellii in both somatic hybrids. In one of the somatic hybrids, 2850, loci on chromosome 1 were also homozygous L. pennellii. The other somatic hybrid, 28F5, was heterozygous at all chromosome 1 loci tested, but exhibited altered stoichiometry of parental bands as compared to the sexual hybrid. Loci on chromosome 2 from both somatic hybrids have altered stoichiometry, with L. pennellii alleles being four times more abundant than expected. Both somatic hybrids contain the L. esculentum chloroplast genome, while only L. pennellii polymorphisms have been detected in the mitochondrial genome.     

Somatic hybridization between Brassica juncea and Moricandia arvensis by protoplast fusion

Intergeneric somatic hybrids have been produced between Brassica juncea (2n=36, AABB) cv. RLM-198 and Moricandia arvensis (2n=28, MM) by protoplast fusion. Hypocotyl protoplasts of B. juncea were fused with mesophyll protoplasts of M. arvensis using polyethylene glycol. Fusion frequency, estimated on the basis of differential morphological characterstics of parental protoplasts was about 5%. Of the 156 calli obtained, four calli produced shoots intermediate in morphology between the parents. Hybrid nature of the plants was confirmed using wheat nuclear rDNA probe. Hybridization of total DNA with a mitochondrial DNA probe carrying 5s–18s rRNA genes of maize showed that the mitochondria of the somatic hybrids were derived from the wild species M. arvensis. Meiosis in the only hybrid that produced normal flowers revealed the occurrence of 64 chromosomes, the sum of chromosomes of parental species. Inspite of complete pollen sterility, siliquas were produced in this hybrid by back-crossing with B. juncea. These siliquas on in vitro culture produced 12 seeds.     

Tomato cybrids with mitochondrial DNA from Lycopersicon pennelli

Summary Cybrids have been regenerated following protoplast fusion of iodoacetamide-treated leaf mesophyll cells of Lycopersion esculentum cv UC82 and gamma-irradiated cell suspensions of L. pennellii, LA716. The cybrids were recovered in the regenerant population at a frequency of 19%, no selection pressure was applied for the persistence of the donor cytoplasm. The nuclear genotype of ten cybrids was characterized extensively using isozyme markers, cDNA-based restriction fragment length polymorphisms (RFLPs), and the morphology of the plants. No nuclear genetic information from L. pennellii was detected in the cybrids. The organellar genotype of the cybrids was determined using cloned probes and species-specific RFLPs. All the cybrids had inherited the tomato chloroplast genome and had varying amounts of L. pennellii mitochondrial DNA. The cybrids all had a diploid chromosome number of 24, produced pollen, and set seed.     

Somatic hybrid plants between Lycopersicon esculentum and Solanum lycopersicoides

Summary Leaf mesophyll protoplasts of Lycopersicon esculentum (2n=2x=24) were fused with suspension culture-derived protoplasts of Solanum lycopersicoides (2n=2x=24) and intergeneric somatic hybrid plants were regenerated following selective conditions. A two phase selection system was based on the inability of S. lycopersicoides protoplasts to divide in culture in modified medium 8E and the partial inhibition of L. esculentum protoplasts by the PEG/DMSO fusion solution. At the p-calli stage, putative hybrids were visually selected based on their hybrid vigor and lime-green coloration in contrast to slower growing parental calli characterized by a watery, whitish-brown coloration. Early identification of the eight hybrid plants studied was facilitated by isozyme analysis of leaf tissue samples taken from plants in vitro at the rooting stage. Regenerated plants growing in planting medium were further verified for hybridity by 5 isozymes marking 7 loci on 5 chromosomes in tomato. These included Skdh-1 mapped to chromosome 1 of tomato, Pgm-2 on chromosome 4, Got-2 and Got-3 on chromosome 7, Got-4 on chromosome 8, and Pgi-1 and Pgdh-2 both on chromosome 12. Fraction I protein small subunits further confirmed the hybrid nature of the plants with bands of both parents expressed in all hybrids. The parental chloroplasts could not be differentiated by the isoelectric points of the large subunit. Seven of the eight somatic hybrids had a chromosome number ranging from the expected 2n=4x=48 to 2n=68. Mixoploid root-tip cells containing 48, 53, 54 or 55 chromosomes for two of the hybrids were also observed.Michigan Agricultural Experiment Station Journal Article No. 11736. Supported by Grant No. I-751-84R from BARD — The United States — Israel Binational Agricultural Research and Development Fund     

Comparison of the organization of the mitochondrial genome in tomato somatic hybrids and cybrids

Summary The organization of the mitochondrial genome in somatic hybrids and cybrids regenerated following fusion of protoplasts from cultivated tomato, Lycopersicon esculentum, and the wild species, L. Pennellii, was compared to assess the role of the nuclear genotype on the inheritance of organellar genomes. No organellar-encoded traits were required for the recorvery of either somatic hybrids or cybrids. The organization of the mitochondrial genome was characterized using Southern hybridization of restriction digestions of total DNA isolated from ten cybrids and ten somatic hybrids. A bank of cosmid clones carrying tomato mitochondrial DNA was used as probes, as well as a putative repeated sequence from L. pennellii mitchondrial DNA. The seven cosmids used to characterize the mitochondrial genomes are predicted to encompass at least 60% of the genome. The frequency of nonparental organizations of the mitochondrial genome was highest with a probe derived from a putative repeat element from the L. pennellii mitochondrial DNA. There was no difference in the average frequency of rearranged mitochondrial sequences in somatic hybrids (12%) versus cybrids (10%), although there were individual cybrids with a very high frequency of novel fragments (30%). The frequency of tomato-specific mtDNA sequences was higher in cybrids (25%) versus somatic hybrids (12%), suggesting a nuclear-cytoplasmic interaction on the inheritance of tomato mitochondrial sequences.     

Organelle analysis of symmetric and asymmetric hybrids between Lycopersicon peruvianum and Lycopersicon esculentum

Summary The organelles of somatic hybrids obtained from symmetric and asymmetric fusions between the Lycopersicon species L. peruvianum and L. esculentum were analyzed by DNA hybridization methods. In the asymmetric fusions the L. peruvianum protoplasts were gamma-irradiated at a dose of 50, 300 and 1,000 Gy. The organelles were characterized using the Petunia chloroplast probe pPCY64 and the mitochondrial EcoRI-SalI fragment of the Pcf gene. In all symmetric and asymmetric hybrid plants, a total of 73 being analyzed, only one of the parental chloroplast genomes was present, except for one hybrid plant which harbored both parental chloroplast genomes. No recombination and/or rearrangement in the chloroplast genome could be identified with the pPCY64 probe. Irradiation of the L. peruvianum protoplasts did not significantly reduce the fraction of asymmetric hybrids with L. peruvianum chloroplasts. A novel mitochondrial restriction pattern was present in 5 out of 24 hybrids tested. In 9 hybrids novel combinations of chloroplasts and mitochondria were found, indicating that both organelle types sorted out independently.     

A rapid procedure for plant regeneration from protoplasts isolated from suspension cultures and leaf mesophyll cells of wild Solanum species and Lycopersicon pennellii

A rapid procedure for protoplast isolation, culture and plant regeneration has been developed for two Solanum species (S. lycoperisicoides and S. verrucosum) and Lycopersicon pennellii. Freshly isolated protoplasts were initially cultured in liquid Solanum Culture Medium (SCM), containing 2,4-dichlorophenoxy acetic acid (2,4-D). Subsequent dilution with fresh culture medium without auxins appeared to be essential to obtain rapid regeneration medium later on. The resulting micro calli were first grown in a culture medium containing 0.5 mg/l 6-BAP and 0.05 mg/l NAA and 0.2 M mannitol and 7.3 mM sucrose to induce greening, at a lower osmolarity (300 mOsm · kg⁻¹). Then, the green micro calli were transferred to shoot induction medium, containing 2 mg/l zeatin, 0.1 mg/l IAA and 2% sucrose (150 mOsm · kg⁻¹). In this way plants could be regenerated from leaf mesophyll protoplasts and suspension cell-derived protoplasts of L. pennellii and S. lycopersicoides within 2 months. Shoot regeneration from leaf mesophyll protoplasts of the two lines of S. verrucosum could be obtained 3 months after protoplast isolation.     

Analysis of nuclear and organellar DNA of somatic hybrid calli and plants between Lycopersicon spp. and Nicotiana spp.

Protoplast fusion experiments between Lycopersicon esculentum or L. peruvianum and Nicotiana tabacum or N. plumbaginifolia were performed to investigate the possibility of producing symmetric and asymmetric somatic hybrids between these genera. These fusions, which involved 1.7 × 10⁸ protoplasts, yielded 35 viable hybrid calli. Plant regeneration was successful with two calli. One of these regenerants flowered, but developed no fruits. Analysis of the nuclear DNA by means of dot blot hybridization with species-specific repetitive DNA probes combined with flow cytometry, revealed that the nuclei of most hybrid calli contained the same absolute amount of Nicotiana DNA as the Nicotiana parent or (much) less, whereas the amount of Lycopersicon DNA per nucleus was 2–5 times that of the parental genotype. Eighteen of the 34 hybrids analyzed possessed Lycopersicon chloroplast DNA (cpDNA), whereas the other 16 had DNA from Nicotiana chloroplasts. The cpDNA type was correlated with the nuclear DNA composition; hybrids with more than 2C Nicotiana nuclear DNA possessed Nicotiana chloroplasts, whereas hybrids with 2C or less Nicotiana nuclear DNA contained Lycopersicon chloroplasts. Mitochondrial DNA (mtDNA) composition was correlated with both nuclear DNA constitution and chloroplast type. Hybrids possessed only or mainly species-specific mtDNA fragments from the parent predominating in the nucleus and often providing the chloroplasts. The data are discussed in relation to somatic incompatibility which could explain the low frequency at which hybrids between Lycopersicon and Nicotiana species are obtained and the limited morphogenetic potential of such hybrids.