Diplotaxis catholica + Brassica juncea somatic hybrids: molecular and cytogenetic characterization

Summary Intergeneric somatic hybrids Diplotaxis catholica (2n=18) + Brassica juncea (2n=36) were produced by fusing mesophyll protoplasts of the former and hypocotyl protoplasts of the latter using polyethylene glycol. Out of 52 somatic embryos, 24 produced plants of intermediate morphology. Cytological analysis of 16 plants indicated that 15 were symmetric hybrids carrying 54 chromosomes, the sum of the parental chromosome numbers. One hybrid was asymmetric with 45 chromosomes. Nuclear hybridity of five putative hybrids was confirmed by the Southern hybridization pattern of full length 18s-25s wheat nuclear rDNA probe which revealed the presence of Hind III fragments characteristic of both the parental species. The hybridization pattern of mitochondria specific gene probe cox I indicated that three of the hybrids carried B. juncea mitochondria and one carried mitochondria of D. catholica. Presence of novel 3.5 kb Hind III and 4.8 kb Bgl II fragments suggested the occurrence of mtDNA recombination in one of the hybrids. The hybrids were pollen sterile. However, seeds were obtained from most of the hybrids by back crossing with B. juncea.     

Interspecific somatic hybrid plants between eggplant (Solanum melongena) and Solanum torvum

Summary Mesophyll protoplasts of eggplant (cv Black Beauty) and of Solanum torvum (both 2n=2x=24) were fused using a modification of the Menczel and Wolfe PEG/DMSO procedure. Protoplasts post-fusion were plated at 1 × 10⁵/ml in modified KM medium, which inhibited division of S. torvum protoplasts. One week prior to shoot regeneration, ten individual calluses had a unique light-green background and were verified as cell hybrids by the presence of the dimer isozyme patterns for phosphoglucoisomerase (PGI) and glutamate oxaloacetate transaminase (GOT). Hybridity was also confirmed at the plant stage by DNA-DNA hybridization to a pea 45S ribosomal RNA gene probe. The ten somatic hybrid plants were established in the greenhouse and exhibited intermediate morphological characteristics such as leaf size and shape, flower size, shape, color and plant stature. Their chromosome number ranged from 46–48 (expected 2n=4x=48) and pollen viability was 5%–70%. In vitro shoots taken from the ten hybrid plants exhibited resistance to a verticillium wilt extract. Total DNA from the ten hybrids was restricted and hybridized with a 5.9 kb Oenothera chloroplast cytochrome f gene probe, a 2.4 kb EcoRI clone encoding mitochondrial cytochrome oxidase subunit II from maize and a 22.1 kb Sal I mitochondrial clone from Nicotiana sylvestris. Southern blot hybridization patterns showed that eight of ten somatic hybrids contained the eggplant cpDNA, while two plants contained the cpDNA hybridization patterns of both parents. The mtDNA analysis revealed the presence of novel bands, loss of some specific parental bands and mixture of specific bands from both parents in the restriction hybridization profiles of the hybrids.Michigan Agricultural Experiment Station Journal Article No. 12545     

Transmission of organelles in triploid hybrids produced by gametosomatic fusions of two Nicotiana species

Summary Gametosomatic hybrids produced by the fusion of microspore protoplasts of Nicotiana tabacum Km⁺Sr⁺ with somatic cell protoplasts of N. rustica were analysed for their organelle composition. For the analysis of mitochondrial (mt)DNA, species-specific patterns were generated by Southern hybridization of restriction endonuclease digests of total DNA and mtDNA with four DNA probes of mitochondrial origin: cytochrome oxidase subunit I, cytochrome oxidase subunit II, 26s rDNA and 5s-18s rDNA. Of the 22 hybrids analyzed, some had parental-type pattern for some probes and novel-type for the others, indicating interaction between mtDNA of the two parent species. For chloroplast (cp)DNA analysis, species-specific patterns were generated by Southern hybridization of restriction endonuclease digests of total DNA with large subunits of ribulose bisphosphate carboxylase and cpDNA as probes. All the hybrids had N. rustica-specific patterns. Hybrids were not resistant to streptomycin, a trait encoded by the chloroplast genome of N. tabacum. In gametosomatic fusions of the two Nicotiana species, mitochondria but not the chloroplasts are transmitted from the parent contributing microspore protoplasts.     

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.     

Organization of nuclear ribosomal DNA and species-specific polymorphism in closely related Fraxinus excelsior and F. oxyphylla

The ribosomal DNA repeat units of two closely related species of the genus Fraxinus, F. excelsior and F. oxyphylla, were characterized. The physical maps were constructed from DNA digested with BamHI, EcoRI, EcoRV and SacI, and hybridized with three heterologous probes. The presence or the absence of an EcoRV restriction site in the 18s RNA gene characterizes two ribosomal DNA unit types found in both species and which coexist in all individuals. A third unit type appeared unique to all individuals of F. oxyphylla. It carries an EcoRI site in the intergenic spacer. Each type of unit displayed length variations. The rDNA unit length of F. excelsior and F. oxyphylla was determined with EcoRV restriction. It varied between 11kb and 14.5kb in F. excelsior and between 11.8kb to 13.8kb in F. oxyphylla. Using SacI restriction, at least ten spacer length variants were observed in F. excelsior, for which a detailed analysis was conducted. Each individual carries 2–4 length variants which vary by a 0.3-kb step multiple. This length variation was assigned to the intergenic spacer. By using the entire rDNA unit of flax as probe in combination with EcoRI restriction, each species can be unambiguously discriminated. The species-specific banding pattern was used to compare trees from a zone of sympatry between the two species. In some cases, a conflicting classification was obtained from morphological analysis and the use of the species-specific rDNA polymorphism. Implications for the genetic management of both species are discussed.     

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.     

Spontaneous extensive chromosome elimination in somatic hybrids between somatically congruent species Nicotiana tabacum L. and Atropa belladonna L.

Summary Mesophyll protoplasts of the kanamycin-resistant nightshade, Atropa belladonna, were fused with mesophyll protoplasts of the phosphinothricin resistant-tobacco, Nicotiana tabacum. A total of 447 colonies resistant to both inhibitors was selected. Most of them regenerated shoots with morphology similar to one of the earlier obtained and described symmetric somatic hybrids Nicotiana + Atropa. However, three colonies (0.2%) regenerated vigorously growing tobacco-like shoots; they readily rooted, and after transfer to soil, developed into normal, fertile plants. Unlike their tobacco parental line, BarD, the obtained plants are resistant to kanamycin [they root normally in the presence of kanamycin (200 mg/1)] and possess activity of neomycin phosphotransferase (NPT II) with the same electrophoretic mobility as the one of the nightshade line. According to Southern blot hybridization analysis carried out with the use of radioactively labeled cloned fragments of the Citrus lemon ribosomal DNA repeat, as well as with Nicotiana plumbaginifolia genus-specific, interspersed repeat Inp, the kanamycin-resistant plants under investigation have only species-specific hybridizing bands from tobacco. Cytological analysis of the chromosome sets shows that plants of all three lines possess 48 large chromosomes similar to Nicotiana tabacum ones (2n = 48), and one small extra chromosome (chromosome fragment) similar to Atropa belladonna ones (2n = 72). Available data allow the conclusion that highly asymmetric, normal fertile somatic hybrids with a whole diploid Nicotiana tabacum genome and only part (not more than 2.8%) of an Atropa belladonna genome have been obtained without any pretreatment of a donor genome, although both these species are somatically congruent.     

Intergeneric hybridization between Pleurotus ostreatus and Schizophyllum commune by PEG-induced protoplast fusion

Intergeneric hybridization between Pleurotus ostreatus and Schizophyllum commune was studied using PEG-induced fusion. The fusion of protoplasts from auxotrophic mutant strains resulted in the formation of fusion hybrids in the frequencies of 3.6 to 7.3×10^–5. Most of these fusion hybrids were monokaryotic and sterile and no heterokaryosis occurred. Most fusants showed a significantly higher nuclear DNA content when compared to parental strains and no diploids (parent 1 genome plus parent 2 genome) were found. Some fusion hybrids revealed both parental fragments in nuclear and mitochondrial rDNA PCR profiles. AP-PCR (Arbitrarily-primed Polymerase Chain Reaction) fingerprints also indicated that most of the fusion products were recombinant hybrids.     

Random chloroplast segregation and frequent mtDNA rearrangements in fertile somatic hybrids between Nicotiana tabacum L. and N. glutinosa L.

Patterns of organelle inheritance were examined among fertile somatic hybrids between allotetraploid Nicotiana tabacum L. (2n=4x=48) and a diploid wild relative N. glutinosa L. (2n=2x=24). Seventy somatic hybrids resistant to methotrexate and kanamycin were recovered following fusion of leaf mesophyll protoplasts of transgenic methotrexate-resistant N. tabacum and kanamycin-resistant N. glutinosa. Evidence for hybridization of nuclear genomes was obtained by analysis of glutamate oxaloacetate transaminase and peroxidase isoenzymes and by restriction fragment length polymorphism (RFLP) analysis using a heterologous nuclear ribosomal DNA probe. Analysis of chloroplast genomes in a population of 41 hybrids revealed a random segregation of chloroplasts since 25 possessed N. glutinosa chloroplasts and 16 possessed N. tabacum chloroplasts. This contrasts with the markedly non-random segregation of plastids in N. tabacum (+)N. rustica and N. tabacum (+) N. debneyi somatic hybrids which we described previously and which were recovered using the same conditions for fusion and selection. The organization of the mitochondrial DNA (mtDNA) in 40 individuals was examined by RFLP analysis with a heterologous cytochrome B gene. Thirty-eight somatic hybrids possessed mitochondrial genomes which were rearranged with respect to the parental genomes, two carried mtDNA similar to N. tabacum, while none had mtDNA identical to N. glutinosa. The somatic hybrids were self-fertile and fertile in backcrosses with the tobacco parent.Contribution No. 1487 Plant Research Centre     

Comparison of the ribosomal RNA genes in four closely relatedCucurbitaceae

Cucurbitaceae are characterized by a high copy number for nuclear ribosomal RNA genes. We have investigated the genomic ribosomal DNA (rDNA) of four closely related species of this family with respect to structure, length heterogeneity, and evolution. InCucumis melo (melon) there are two main length variants of rDNA repeats with 10.7 and 10.55kb.Cucumis sativus (cucumber) shows at least three repeat types with 11.5, 10.5, and 10.2kb.Cucurbita pepo (zucchini) has two different repeat types with 10.0 and 9.3kb. There are also two different repeat types inCucurbita maxima (pumpkin) of about 11.2 and 10.5kb. Restriction enzyme mapping of the genomic rDNA of these four plants and of cloned repeats ofC. sativus shows further heterogeneities which are due to methylation or point mutations. By comparison of the restriction enzyme maps it was possible to trace some evolutionary events in the family ofCucurbitaceae. Some aspects of regulation and function of the middle repetitive rRNA genes (here between 2000 and 10000 copies) are discussed.     

Production and characterization of fertile somatic hybrids of eggplant (Solanum melongena L.) with Solanum aethiopicum L.

Summary In order to produce fertile somatic hybrids, mesophyll protoplasts from eggplant were electrofused with those from one of its close related species, Solanum aethiopicum L. Aculeatum group. On the basis of differences in the cultural behavior of the parental and hybrid protoplasts, 35 somatic hybrid plants were recovered from 85 selected calli. When taken to maturity either in the greenhouse or in the field, the hybrid plants were vigorous, all rapidly overtopping parental individuals. The putative hybrids were intermediate with respect to morphological traits, and all of their organs were larger, particularly the leaves and stems. DNA analysis of the hybrids using flow cytometry in combination with cytological analysis showed that 32 were tetraploids, 1 hexaploid and 2 mixoploids. The hybrid nature of the 35 selected plants was confirmed by a comparison of the isoenzyme patterns of isocitrate dehydrogenase (Idh), 6-phosphogluconate dehydrogenase (6-Pgd) and phosphoglucomutase (Pgm). Chloroplast DNA (ctDNA) restriction analysis using Bam HI revealed that among the 27 hybrid plants analyzed, 10 had S. aethiopicum patterns and the 17 remaining hybrids exhibited bands identical with those of eggplant without any changes. All of the somatic hybrid plants flowered. Both parental plants had 94% stainable pollen, while the hybrids varied widely in pollen viability ranging from 30% to 85%. The somatic hybrids showed high significant variation in fruit production. Nevertheless, there was a tendency for low fertility to be associated often with S. aethiopicum chloroplast type and/or with an abnormal ploidy level, while good fertility was mostly associated with the tetraploid level and eggplant chloroplasts. Interestingly, 2 tetraploid somatic hybrid clones were among the most productive, yielding up to 9 kg/plant. As far as the fertility of the F₁ sexual counterpart was concerned, only 2 fruits of 50 g were obtained. Hybrid fertility in relation to phylogenetic affinities of the fusion partners is discussed.     

Differential fate of plastid and mitochondrial genomes in Petunia somatic hybrids

Summary The chloroplast (cp) and mitochondrial (mt) DNAs of Petunia somatic hybrid plants, which were derived from the fusion of wild-type P. parodii protoplasts with albino P. inflata protoplasts, were analyzed by endonuclease restriction and Southern blot hybridization. Using ³²P-labelled probes that distinguished the two parental cpDNAs at a BamH1 site and at a HpaII site, only the P. parodii chloroplast genome was detected in the 10 somatic hybrid plants analyzed. To examine whether cytoplasmic mixing had resulted in rearrangement of the mitochondrial genome in the somatic hybrids, restriction patterns of purified somatic hybrid and parental mtDNAs were analyzed. Approximately 87% of those restriction fragments which distinguish the two parental genomes are P. inflata-specific. Restriction patterns of the somatic hybrid mtDNAs differ both from the parental patterns and from each other, suggesting that an interaction occurred between the parental mitochondrial genomes in the somatic fusion products which resulted in generation of the novel mtDNA patterns. Southern blot hybridization substantiates this conclusion. In addition, somatic hybrid lines derived from the same fusion product were observed to differ in mtDNA restriction pattern, reflecting a differential sorting-out of mitochondrial genomes at the time the plants were regenerated.     

Development of Molecular Identification Method for Genus Alexandrium (Dinophyceae) Using Whole-Cell FISH

We have developed a method to identify species in the genus Alexandrium using whole-cell fluorescent in situ hybridization with FITC-labeled oligonucleotide probes that target large subunit ribosomal rRNA molecules. The probes were designed based on the sequence of the rDNA D1-D2 region of Alexandrium species. DNA probes specific for toxic A. tamarense and A. catenella and nontoxic A. affine, A. fraterculus, A. insuetum, and A. pseudogonyaulax, respectively, were applied to vegetative cells of all above Alexandrium species to test the sensitivity of the probes. Each DNA probe hybridized specifically with vegetative cells of the corresponding Alexandrium species and showed no cross-reactivity to noncorresponding Alexandrium species. In addition, no cross-reactivity of the probes was observed in experiments using concentrated natural seawater samples. The TAMAD2 probe, which is highly specific to A. tamarense, a common toxic species in Korean coastal waters, provides a simple and reliable molecular tool for identification of toxic Alexandrium species.     

Ribosomal RNA genes in species of theCynareae tribe (Compositae). II

Summary The structure of the ribosomal DNA has been analyzed in three species of theCynareae tribe using Southern blot hybridization.Silybum marianum possesses two types of ribosomal genes 12.3 and 13.4 kb long;Cirsium vulgare has at least four types of rDNA repeats 10.6, 10.5, 11.7 and 11.9 kb long;Carlina acaulis only one type of ribosomal genes 9.7 kb long. The rRNA genes of the three species studied showed an identical restriction mapping in the 18 S and 25 S regions. However species differentiation in length and/or nucleotide sequences are present in the external spacer and very probably in the internal transcribed spacer.By cytophotometric studies and byin vitro rRNA/DNA filter hybridization, the DNA amount/4 C nucleus and the rDNA percentage were calculated in nine species of theCynareae tribe:Cynara cardunculus subsp.scolymus (artichoke),Cynara cardunculus subsp.cardunculus (wild artichoke),Onopordon acanthium, O. illyricum, Galactites tomentosa, Carduus nutans, Silybum marianum, Cirsium vulgare andCarlina acaulis. The DNA amount/4 C nucleus in eight species are similar, ranging from 4.24 pg inGalactites tomentosa to 5.96 pg inCirsium vulgare, whileCarlina acaulis has a DNA amount/ 4C nucleus of 11.8 pg. The rDNA percentages range from 0.192% inOnopordon acanthium to 1.022% inSilybum marianum, whileCarlina acaulis has 0.038% of rDNA. This latter species is clearly distinct within theCynareae tribe.     

The fate of ribosomal genes in three interspecific somatic hybrids of Medicago sativa: three different outcomes including the rapid amplification of new spacer-length variants

We have characterized the genetic consequences of somatic hybridization within the ribosomal DNA (rDNA) of three interspecific hybrids, each involving M. sativa as one of the parents. Restriction-fragment-length-polymorphisms (RFLPs) of rDNA spacers and fluorescent-in-situ-hybridization (FISH) of an 18S-gene probe to mitotic chromosomes were used to compare parental and hybrid species. The M. sativa-coerulea hybrid retained all six parental nucleolar-organizing regions (NORs) and all parental RFLPs representing a complete integration of rDNA. The M. sativa-arborea hybrid retained five of six parental NORs while losing half of the arborea-specific RFLPs, indicating that simple chromosome loss of one arborea NOR accounted for the RFLP losses. Dramatic alterations occurred within the M. sativa-falcata hybrid where five of six parental NORs were retained and new rDNA RFLPs were created and amplified differentially among somaclonal-variant plants. The molecular basis of the new RFLPs involved increased numbers of a 340-bp subrepeating element within the rDNA intergenic spacer (IGS), suggesting that recurrent cycles of unequal recombination occurred at high frequency within the rDNA in somatic lineages.This paper was supported by the National Research Council of Italy, Special Project RAISA, Sub-project No. 2, Paper No. 1077     

Relative rates of divergence of spacer and gene sequences within the rDNA region of species in the Triticeae: Implications for the maintenance of homogeneity of a repeated gene family

Summary The relative rates of divergence of 11 regions of the wheat rDNA cloned in pTA250 were estimated by measuring sequence change in 6 Triticum species. The Tm analysis of ³²P probes synthesized from the pTA250 regions and hybridized to DNA from the Triticum species provided an estimate of sequence change relative to T. aestivum. The results revealed a region of 1.2 kb preceding the 18S rRNA gene which was more conserved than the rest of the spacer. In addition the transcribed spacer between the 18S and 26S rRNA genes was shown to be poorly-conserved; the genes, as expected, were highly conserved. A model which proposes RNA as a co-factor in gene conversion is suggested to account for the observations.     

Morphological and molecular characterization of somatic hybrid plants between Lycopersicon esculentum and Solanum nigrum

Summary Mesophyll protoplasts of tomato (Lycopersicon esculentum Mill. var. cerasiforme) and of an atrazine-resistant biotype of black nightshade, (Solanum nigrum L.), were fused by using polyethylene glycol/dimethyl sulfoxide (PEG/DMSO) solution and three somatic hybrid plants, each derived from a separate callus, were recovered. A twostep selection system was used: (1) protoplast culture medium (modified 8E) in which only tomato protoplasts formed calluses; and (2) regeneration medium (MS2Z) on which only S. nigrum calluses produced shoots. These selective steps were augmented by early isozyme analysis of putative hybrid shoots still in vitro. Phosphoglucoisomerase (PGI) and glutamate oxaloacetate transaminase (GOT) mapped to five loci on four chromosomes in tomato confirmed the hybrid nature of the nuclei of regenerated shoots. The somatic hybrid plants had simple leaves, and intermediate flower and bud morphology, but anthesis was reduced to 5% due to premature bud abscission and the pollen grains were non-viable. Southern DNA blot hybridization using a pea 45 S ribosomal RNA gene probe reconfirmed the hybrid nature of the nuclear genome of the three plants. A ³²P-labeled probe of Oenothera chloroplast DNA (cpDNA) hybridized to cpDNA restricted with EcoRI or EcoRV indicated the presence of the tomato cpDNA pattern in all three hybrids. Likewise, the plants were all found to be atrazine sensitive. Analysis with two mitochondrial (mt)DNA-specific probes, maize cytochrome oxidase subunit II and PmtSylSa8 from Nicotiana sylvestris, showed that, in addition to typical mitochondrial rearrangements, specific bands of both parents were present or missing in each somatic hybrid plant.Michigan Agricultural Experiment Station Journal Article No. 12433     

Purification and restriction endonuclease mapping of soybean 18 S and 25 S ribosomal RNA genes

The restriction endonuclease map of the 25 S and 18 S ribosomal RNA genes of a higher plant is presented. Soybean (Glycine max) rDNA was enriched by preparative buoyant density centrifugation in CsCl-actinomycin D gradients. The buoyant density of the rDNA was determined to be 1.6988 g cm^–3 by analytical centrifugation in CsCl. Saturation hybridization showed that 0.1% of the total DNA contains 25 S and 18 S rRNA coding sequences. This is equivalent to 800 rRNA genes per haploid genome (DNA content: 1.29 pg) or 3200 for the tetraploid genome. Restriction endonuclease mapping was performed with Bam H I, Hind III, Eco R I, and BstI. The repeating unit of the soybean ribosomal DNA has a molecular weight of 5.9·10⁶ or approximately 9,000 kb. The 25 S and 18 S rRNA coding sequences were localized within the restriction map of the repeating unit by specific hybridization with either [¹²⁵I]25 S or [¹²⁵I]18 S rRNA. It was demonstrated that there is no heterogeneity even in the spacer region of the soybean rDNA.     

Ribosomal RNA genes in species of theCynareae tribe (Compositae). I.

Summary By means of Southern blot hybridization, the structure of the ribosomal DNA in six species of theCynareae tribe has been analyzed. Artichoke and wild artichoke possess only one type of ribosomal genes 13 kb long;Onopordum acanthium has at least two types of rDNA repeats 9.9 kb and 10.3 kb long andO. illyricum has a third gene type 9.7 kb long; inGalactites tomentosa there are at least four ribosomal gene types of 10.9, 11.6, 11.5, and 10kb;Carduus nutans possesses two ribosomal gene types of the same length of 12.5 kb that vary in the nucleotide sequence of the external spacer. The rRNA genes of all the species studied have an identical restriction mapping in the 18 S and 25 S regions, differences in length and/or nucleotide sequence are present in the external spacer.     

Rearrangements in the mitochondrial genome of somatic hybrid cell lines of Pennisetum americanum (L.) K. Schum. + Panicum maximum Jacq.

Summary Mitochondrial DNA from three somatic hybrid cell lines of Pennisetum americanum + Panicum maximum was compared with mitochondrial DNA of the parents. Gel electrophoresis of BamHI-restricted mitochondrial DNA indicated that extensive rearrangements had occurred in each of the three hybrid lines. The hybrid restriction patterns showed a combination of some bands from each parent plus novel fragments not present in either parent. Additional evidence for rearrangements was obtained by hybridization of eight DNA probes, carrying sequences of known coding regions, to Southern blots. Each of the somatic hybrids exhibited a partial combination of the parental mitochondrial genomes. These data suggest recombination or amplification of the mitochondrial genomes in the somatic hybrids.