ABSTRACT
Several hybrid callus lines were produced through somatic hybridization between the diploid transformed Solanum tuberosum plant clone 413 (2n = 2x = 24) and a diploid wild-type plant clone of Nicotiana plumbaginifolia (2n = 2x = 20). The hybrid callus lines with subdiploid numbers of potato chromosomes were studied for karyotypic evolution as well as for segregation of the transformation marker characters (i.e. hormone autotrophy, opine synthesis, kanamycin resistance and ß-glucuronidase activity). Initially, these hybrids (cultured in kanamycin-containing medium) expressed all of the transformation characters. Six callus lines were selected for the establishment of cell suspension cultures; two of these were also used to initiate sublines, one from single cells of a suspension culture, and the other from callus-derived protoplasts. The cell suspension cultures and the sublines were cultured in kanamycin-free medium. After prolonged culture, karyotypic analysis of the various cell suspension lines revealed independent evolution of both parental genomes. Out of the six suspension lines, four showed a considerably reduced number of potato chromosomes as compared to the original hybrid callus lines, whereas the karyotypes of the individual sublines generally reflected the karyotypic diversity of the original cultures. The fate of the marker characters in various suspension cultures and sublines revealed independent segregation of the markers of TL-DNA (hormone autotrophy) and vector T-DNA (kanamycin resistance and ß-glucuronidase activity). Loss of the TR-DNA marker (opine synthesis) was observed only in combination with the simultaneous loss of the TL-DNA marker and the vector T-DNA markers. The results on segregation patterns of marker characters are discussed in the light of specific chromosome loss in the hybrid lines and gene linkage relationships.
ABSTRACT
Electrofusion was carried out between mesophyll protoplasts from the transformed diploid S. tuberosum clone 413 (2n=2x=24) which contains various genetic markers (hormone autotrophy, opine synthesis, kanamycin resistance, ß-glucuronidase activity) and mesophyll protoplasts of a diploid wild-type clone of N. plumbaginifolia (2n=2x=20). Hybrid calli were obtained after continuous culture on selection medium containing kanamycin. Parental chromosome numbers, determined at 2 months after fusion, revealed hybrid-specific differences between the individual calli. On the basis of these differences three categories of hybrids were distinguished. Category I hybrids contained between 8 and 24 potato chromosomes and more than 20 N. plumbaginifolia chromosomes; category II hybrids had between 1 and 20 N. plumbaginifolia chromosomes and more than 24 potato chromosomes; category III hybrids contained diploid or subdiploid numbers of chromosomes from both parents. The hybrids were evenly distributed over the three categories. After a 1-year culture of 24 representative hybrid callus lines on selection medium the karyotype of 10 hybrids remained stable, whereas 8 hybrids showed polyploidization of the genome of one parent, together with no or minor changes of the chromosome numbers of the other parent. Six hybrids showed slight changes in the hybrid karyotype. The elimination of chromosomes of a particular parent was not correlated to their metaphase location. The processes of spontaneous biparental chromosome elimination leading to the production of asymmetric hybrids of different categories are discussed.
ABSTRACT
The effects of the spindle toxins colchicine, oryzalin and amiprophos-methyl (APM) on metaphase arrest, chromosome scattering, and on the induction and yield of micronuclei were compared in suspension cells ofNicotiana plumbaginifolia (kanamycin-resistant "Doba" line). The inhibition of spindle formation is stronger with oryzalin and APM than with colchicine, which resulted in a more efficient accumulation of meta-phases with well-scattered chromosomes, allowing the isolation of single chromosomes. Further, APM and oryzalin treatments resulted in a higher frequency of micro-nucleated cells and greater yield of micronuclei than after colchicine treatment. The different actions of the chemicals on the functioning of the spindle, development of nuclear membranes around the chromosomes, formation of micronuclei and fusion of micronuclei, resulting in restitution nuclei, are discussed.
ABSTRACT
Agrobacterium transformation of stem internodes of four monohaploid (839-79, 849-7, 851-23, 855-1) and two diploid (M9 and HH260) potato genotypes using hairy root-inducing single (LBA 1020, LBA 9365, LBA 9402) and binary (LBA 1060KG) vectors is reported. Various media and successive culture steps were tested for plant regeneration from different transformed root clones. The fate of introduced genetic markers in root clones and regenerated plants (hairy root phenotype, hormone autotrophy, opine production, kanamycin resistance, ß-glucuronidase activity), the ploidy stability and protoplast yield were analysed. The transformation efficiency of stem internodes (hairy root production) and the regeneration capacity of the transformed root clones greatly differed within and between the various potato genotypes. The regenerated plants obtained after transformation with both types of vectors often showed the absence of one or more genetic markers. However, transformation with the binary Agrobacterium vector generally resulted in the stable presence of the opines in all transformed root clones and most regenerated plants. In HH260, transformation efficiency, plant regeneration of transformed root clones, protoplast yield and ploidy stability were the highest as compared to the other genotypes. The application of these transformed plants as marker lines in gene mapping and gene expression studies is indicated.
ABSTRACT
Flow cytometry and karyological analysis were used to study polysomaty and polyploidization during the first 15 days of callus formation in leaf segments from shoot cultures and greenhouse-grown plants of various lines and genotypes of Solanum tuberosum and S. phureja. The greenhouse-grown plants showed a higher degree of polysomaty (77% and 49% of polyploidized nuclei) than the shoot cultures (< 3%). During the in vitro culture period, polyploidization occurred through endoreduplication. Segments of the five shoot cultures showed up to 87%, 53%, 59%, 45% and 56% polyploidization, respectively; the DNA content of corresponding interphase nuclei amounted to 8C, 16C, 16C, 16C and 8C, and the chromosome numbers to 96. Segments from the two greenhouse-grown genotypes showed up to 87% and 84% polyploidization; the DNA content amounted to 32C and 16C, and the chromosome numbers to 192 and 96. The number of reduplication cycles was species-dependent; the degree of polyploidization was dependent on the initial ploidy level of the genotypes. Cell proliferation did not take place at a constant rate. The maximum frequencies of metaphases (52-171 per segment) occurred after 1 week of culture and were correlated with the ploidy level of the genotypes. Cells were triggered to mitosis rather than to endoreduplication. Cell cycles with normal monochromosomes could be shorter than 1 day, and those with diplochromosomes lasted at least 1 day. Polysomaty, degree of polyploidization and abnormal nuclear processes are discussed in relation to the origin of genetic instability early in culture.
ABSTRACT
Amiprophos-methyl (APM) is a potential herbicide which acts at the level of microtubules. By exposure of suspension cells of Nicotiana plumbaginifolia to this agent, a high degree of metaphase arrest was observed and single as well as groups of chromosomes were scattered throughout the cell, offering good prospects for application in cytology and chromosome isolation. After prolonged exposure to the drug, the chromosomes decondensed and micronuclei were formed. Based on their DNA content, the micronuclei were sorted by flow cytometry. Prospects for application of isolated micronuclei for partial genome transfer and gene mapping are discussed.
ABSTRACT
Attempts were made to obtain plant regeneration in vitro from leaf tissue and stem internodes of Haplopappus gracilis. The morphogenetic effects of several combinations of auxin/kinetin and light with three types of basal media were investigated. A modified White's medium supplemented with IAA gave optimum callus growth, and permitted root and shoot differentiation.
ABSTRACT
A wide range of phenotypic variation occurred among protoplast - derived plants of tetraploid potato cultivar 'Bintje'. The variant plants had alterations in growth and vigour, and in leaf and stem characteristics. The results suggest that the altered morphologies are caused predominantly by changes in ploidy levels. Some alterations could be attributed typically to octoploidy and aneuploidy. The occurrence of mixoploidy indicates that at least part of the observed variation arose during culture stage. The exogeneous cytokinin or auxin level and their combination during in vitro phase influenced the frequency of the variants observed. The origin of variation is discussed.
ABSTRACT
The transmission rate of trisomy was determined for two primary trisomic types, triplo-1 and triplo-3, of the self-incompatible species Lycopersicum peruvianum. Chromosome counts in somatic metaphases of root-tip squashes from 112 progeny plants showed that 8 individuals (7.2 %) were trisomic and 104 (92.8%) were diploid. The average frequency of transmission approximated 2.6% in triplo-1 and 8.6% in triplo-3. Data are presented on the karyotype and the morphological features of the 8 trisomics detected in the progenies of triplo-1 and triplo-3 and the various factors affecting the transmission rate of trisomy are discussed.The transmission rate of trisomy was also determined for the trisomic plant 269 which displayed a complete deletion of the satellited part of chromosome 2 and was characterized by ovate fruits. Out of 18 progeny plants analysed, 8 (44.4%) were trisomic and 10 (55.6%) were diploid. Cytological and morphological analyses of the 8 trisomic individuals revealed that only two of them (11.1 %) resembled the parental trisomic. A number of diploid and trisomic progenies exhibited a partial or a complete deletion of the satellited segment of chromosome 2.
ABSTRACT
Gamma ray treatments at various doses were applied to flowers after cross-compatible pollinations (S1S4 × S12S13) and self-incompatible pollinations (S1S4 × S1S4). After 200 kR treatment (highest dose) a high percentage of pollen became non-viable, and tube growth of all the germinated pollen was inhibited at the first third, or just before half the length of the style. Observations on the ultrastructural aspects revealed that the pollen tubes were destroyed by a precise degradation process which led to the disappearance of the inner wall, lysis of the tube, accumulation of several bipartite particles and alteration of endoplasmic reticulum into a whorl of concentric circles. These results indicated that the high dose of gamma rays probably interferes with protein synthesis. The ultrastructural aspects of compatible pollen tubes after gamma irradiation were similar to those of self-incompatible tubes and therefore it is suggested that cessation of protein synthesis might also be a result of incompatibility.Electron microscopy observations after treatment with 200 kR gamma rays on flowers after self-incompatible pollination (S1S4 × S1S4) showed that gamma irradiation affects the self-incompatibility reaction; but the results obtained so far are difficult to explain.
ABSTRACT
An attempt was carried out to produce trisomics of the wild tomato L. peruvianum, to define their essential features, and to detect relationships between trisomy and the expression of self-compatibility.Triploid-diploid crosses in L. peruvianum yielded nearly 40% aneuploids. Of these, 18% were single trisomics, and the rest had 2, 3 and 4 extra chromosomes. Almost all the trisomics occurred in crosses where the triploid was used as female parent. Vigour and fertility of trisomics were not much different from those of disomics, and morphologically they were very similar.The extra chromosome was identified in three self-compatible trisomic plants through somatic and pachytene chromosome morphology. One of these plants was trisomic for chromosome 1, while the other two were trisomic for chromosome 3. In these trisomics a positive correlation was found between chromosome length and trivalent formation, but no relationship between chromosome length and frequency of laggards was observed.A series of test-crosses revealed that the capacity of the trisomics to produce seed upon selfing always resulted from alterations of the incompatibility phenotype of the style and not from competitive interaction in the pollen. Progeny analyses showed that the self-compatibility features of the trisomics were not transmitted from one generation to the next. The implications of these findings are discussed.
ABSTRACT
An investigation was carried out of the effect of X-irradiation on the size of the mutated sector in three diploids (A.S.D.8, T.N. 1 and G.E.B. 24) and the autotetraploid of G.E.B 24 of Oryza sativa L., by analyzing the M2 segregation ratios of chlorophyll mutants. A total of 612 segregating M1 panicle progenies in the diploids and 284 panicle progenies in the tetraploid was studied in the M2 generation for the ratio of normal to chlorophyll mutant seedlings. Of the 573 segregating M1 panicles analysed by chi square test in the diploids, 206 showed good fit with a 3â¶1 ratio, while the rest of the progenies (367) deviated significantly in both directions from the expected 3â¶1 segregation. This was interpreted as the result of M1 panicles developing from both single cell and multicellular initials following X-irradiation, the former segregating for 3â¶1, the latter significantly deviating from 3â¶1. An assessment of fertility (as judged from the number of M2 seedlings per mutated M1 panicle), related to the distribution of chi square deviations from the 3â¶1 ratio in the diploids, indicated that the M1 panicle progenies deriving from single cell initials had low fertility but those originating from multicellular initials ranged from low to high fertility. A comparison of the distribution pattern of the M2 segregation ratios of chlorophyll mutant phenotypes in the diploid and autotetraploid G.E.B. 24 indicated that, in the tetraploid, a larger sector and only one, or fewer cell initials than in the diploid species, are involved.
ABSTRACT
A comparative study on the relative mutagenicity of radiations (X-and gamma rays) and chemical mutagens (EMS, MMS, dES and NEU) in inducing chlorophyll, viable and total mutations was made on three cultivated varieties (Co. 11, Co. 12 and Co. 18) of Eu-Sorghum. A critical comparison of the chlorophyll, viable and total mutations shows that the mutation rate increased with an increase in dose up to a certain dose level beyond which the saturation effect was observed. EMS was found to be the most potent mutagen in inducing chlorophyll, viable and total mutations. The treatments with NEU (viable mutations) and dES (viable and total mutations) also proved to be effective. In a large number of cases the relationship between chlorophyll and total mutations was more close than that of viable to total mutation rates.
