Majority of random cDNA clones correspond to single loci in the tomato genome

Summary The number of loci corresponding to each of 34 unique, random cDNA clones has been determined for the diploid plant species Lycopersicon esculentum (tomato) (2n=24). Fifty-three percent of the clones are homologous to loci represented only once in the tomato chromosomes. Thirty-two percent of the clones correspond to two genetically independent loci. The remaining clones belong to gene families represented by 3–5 loci. To determine the number of gene copies per locus, reconstruction experiments were performed on six of the single locus clones. The majority of these loci were estimated to contain only 1 or 2 copies of the gene. These results support the concept that the majority of structural genes in this diploid plant species are arranged in single loci and present in low copy number. Multigene families, such as those for the small subunit of ribulose bisphosphate carboxylase, chlorophyll a/b binding polypeptide and actin are exceptions to this rule.     

Plant regeneration from cultured leaf explants of eight wild tomato species and two relatedSolanum species

Summary Optimum conditions for plant regeneration from cultured leaf explants were ascertained for eight different wild tomato (Lycopersicon) species and two closely relatedSolanum species. Of the eight media tested, basal MS medium supplemented with 5 μM of the cytokinin 6-benzyladenine proved to be the best overall regeneration medium. Regeneration frequency varied significantly between species with maximum frequency of regeneration observed forL. chilense, L. peruvianum, andS. lycopersicoides.     

Endosperm dosage relationships among Lycopersicon species

Summary Accessions of eight Lycopersicon species and five yellow-flowered Solanum species were used as males in crosses with 2x and 4x L. esculentum to observe seed set and progeny ploidy. Species which failed in crosses to L. esculentum were crossed as males to 2x and 4x L. peruvianum. In cases of low seed set, chromosome counts were undertaken to establish the nature of the progeny. Endosperm Balance Number (EBN) relationships were determined for the crossability groups. Results support the basic concept of an L. esculentum crossability complex and an L. peruvianum crossability complex. Within the L. esculentum complex, all EBNs appear identical with a value of 2. Within the L. peruvianum complex, more variability appears to exist. The EBN values of this group are higher, and may be approximately double those of the L. esculentum complex. The EBN of L. peruvianum var humifusum appears to be somewhat lower than other L. peruvianum types. The EBN values of S. lycopersicoides, S. rickii, S. ochranthum and S. juglandtfolium could not be determined experimentally. Differential aspects of Lycopersicon and tuber-bearing Solanum evolution may be interpreted on the basis of endosperm compatibility.Co-operative investigation of the Vegetable Crops Research Unit, U.S. Department of Agriculture, Agricultural Research Service, and the Wisconsin Agricultural Experiment Station     

A genetic analysis of cell culture traits in tomato

Summary Tomato genotypes superior in regenerating plants from protoplast and callus cultures were obtained by transferring regeneration capacity from Lycopersicon peruvianum into L. esculentum by classical breeding. The genetics of regeneration and callus growth have been studied in selfed and backcross progenies of a selected plant (MsK93) which has 25% L. peruvianum in its ancestry. Segregation data showed that the favourable cell culture traits of L. peruvianum are dominant. Regeneration capacity from established callus cultures was controlled by two dominant genes. Callus growth on primary expiants, callus growth of established cultures and shoot regeneration from explants had high heritabilities (0.47, 0.78, 0.87, respectively). Callus growth and regeneration capacity were not correlated within the populations studied.     

Euploidy in Ricinus. I. Euploidy and gene dosage effects on cellular proteins

Investigation of an euploid series of castor bean, Ricinus communis L., consisting of haploid, diploid, and tetraploid individuals, was performed to determine the value of such a series in studying the biochemical consequences of genome multiplication. The effects of euploidization of the nuclear genome on the biosynthesis of cellular proteins were examined. Extracts of total soluble proteins from 10-day-old leaves of all three ploidy levels examined by electrophoresis on polyacrylamide gels revealed no difference in the complement of proteins present; however, differences in intensity of several protein bands were detected. Analysis of esterase isozyme activity by isoelectric focusing revealed both increases and decreases in the activity levels of individual isozyme variants in response to changes in ploidy levels. Results from this analysis are discussed in terms of possible regulatory mechanisms active in the regulation of duplicated genes.     

Pollen irradiation in tomato: minor effects on enzymic gene transfer

Summary Pollen irradiation was tested as a possible means of expediting gene transfer in tomato breeding. The experiment was also designed to examine the possibility that irradiation effects which have been previously reported for Nicotiana were due to gametophytic selection against mutated male and female gametes.Pollen from the wild tomato species Solanum pennellii was irradiated with -rays and used in crosses with Lycopersicon esculentum. Pollen from five M1 hybrid plants and five control hybrids was used in backcrosses to an esculentum variety and the same plant also functioned as female parents in crosses with normal S. pennellii pollen. Seven enzymic gene markers which differ between the species were assayed in the backcross populations in the two directions. Allele frequencies did not differ between the populations that were derived from the M1 or F1 hybrids except for one marker, Pgm-2. Elimination of male and female gametes containing the irradiated parent allele of Pgm-2 was observed in a single population. This gene is located on chromosome 4 close to the gamete eliminator (Ge) locus. A review of some of the main characteristics of Ge raises the possibility of the existence of a controlling element activity.     

Different regeneration potential of mesophyll protoplasts from cultivated and a wild species of tomato

Mesophyll protoplasts were isolated from leaves of three cultivars of Lycopersicon esculentum (L.) Mill., namely Hilda 72, Rutgers and Rentita, and from the wild tomato species Lycopersicon peruvianum (L.) Mill. Protoplasts from L. peruvianum divided and grew actively in a liquid medium according to Zapata et al. (1977), whereas protoplasts from the tomato cultivars Hilda 72 and Rutgers showed comparable rates for cell division only, when the content of major elements in this medium was reduced to one half of the original concentration and when mannitol as osmoticum was replaced by glucose. In Rentita protoplasts no cell division could be observed in about 15 different modifications of the five basic culture media tested. The morphogenetic potential of these tomato cells was assessed by comparing the root and shoot formation of protoplasts and of leaf explants. L. peruvianum exhibited the highest potential. Calli derived from protoplasts regenerated roots on Murastrige-Skoog agar containing 1 M benzylaminopurine (BAP) plus 10 M indole-3-acetic acid (IAA) and 0.1 M BAP plus 1 M IAA. Shoot formation occurred in the combinations of 10 M BAP with 0.1, 1.0, and 10 M IAA. Plantlets regenerated from the L. peruvianum calli could be grown in soil. No shoots or roots were regenerated from calli of Hilda 72 and Rutgers protoplasts in all combinations of BAP and IAA tested in the range from 0.1 M to 100 M, thus indicating the rather low morphogenetic potential of these protoplasts as compared to protoplasts from L. peruvianum leaves.Abbreviations BAP benzylaminopurine - IAA indole-acetic acid - TMV tobacco mosaic virus     

Genetic variation inSolanum pennellii: Comparisons with two other sympatric tomato species

Genetic variation—primarily in 19 genetic loci of seven enzyme systems—was analyzed in accessions from various parts of the geographic range ofSolanum pennellii, which according to all tested biosystematic criteria behaves like a species ofLycopersicon. In comparison with the largely sympatricL. hirsutum andL. pimpinellifolium, this species exhibits the same trends of reduced allogamy and decreased genetic variation toward the north and south margins of its distribution, though to a much lesser degree; it does not exhibit their trends toward smaller flower size in the same peripheral regions. All three species agree to a considerable extent in the ranking of their tested loci in respect to degree of variablility; however, overall polymorphy is highest inS. pennellii. Except for the appearance of self-compatibility at its southernmost margin,S. pennellii is exclusively and rigidly self-incompatible. Alleles are distributed much more uniformly over the range than in the previously mentioned species, marginal and internal endemic mutants being much less abundant. A marked geographic disagreement is evident in regions of high and low variation. These differences in patterns of genetic variability are reconciled in terms of observed differences in mating systems, probable age of distributions, and adaptive strategies.     

Low temperature effect on selective fertilization by pollen mixtures of wild and cultivated tomato species

Summary In vitro pollen germination of cultivated tomato, Lycopersicon esculentum Mill., is inhibited by an ambient temperature of 5°C, more so than pollen from a Peruvian ecotype of Lycopersicon hirsutum Humb. & Bonpl. originating from an altitude of 3200 m. The frequency of L. hirsutum gametes contributing to hybrid zygote formation is more than doubled when controlled fertilizations with pollen mixtures of the two species occurs at 12/6°C as compared to crosses with the same mixtures at 24/19°C. The results suggest that differential selection at the gametophytic level occurs in response to low temperature regimes. To our knowledge this is the first time in higher plants that alteration of an environmental factor has been demonstrated to change selection values of male gametophytes in a fashion predicted by the ecology of the parental sporophytes.     

Shoot regeneration capacity from roots and transgenic hairy roots of tomato cultivars and wild related species

The organogenetic competence of roots and Agrobacterium rhizogenes-induced hairy roots of twelve Lycopersicon genotypes was investigated. Both roots and hairy roots of L. peruvianum, L. chilense, L. hirsutum and two L. peruvianum-derived genotypes regenerated shoots after 2–4 weeks of incubation on zeatin-contained medium. Anatomical analysis showed that shoot regeneration in roots could be direct or indirect, depending on the genotype considered. Hairy roots showed considerable differences in their morphogenetic responses, when compared to the corresponding non-transgenic roots. The differences observed may reflect the influence of the introduced rol genes on hormonal metabolism/sensitivity. Hairy root-derived T0 plants had shortened internodes, wrinkled leaves and abundant root initiation, and most produced flowers and fruits with viable seeds. The hairy root syndrome was detected early in germinating T1 seedlings as a strong reduction in the hypocotyl length. Our data point to the possibility of the use of A. rhizogenes, combined with regenerating Lycopersicon genotypes, in a very simple protocol, based on genetic capacity instead of special procedures for regeneration, to produce transgenic tomato plants expressing rol genes, as well as, genes present in binary vectors. Furthermore, the regeneration differences observed in each Lycopersicon genotype and in transgenic materials expressing rol genes open the possibility for their use in the analysis of both the biochemical and the genetic background of organogenetic competence. This revised version was published online in June 2006 with corrections to the Cover Date.     

Response to chilling of tomato mesophyll protoplasts

Freshly isolated protoplasts from tomato leaves show two completely different responses to a chilling treatment of 12 h at 7° C prior to culture at 29° C, depending on the presence or absence of glucose in the medium. In the culture medium with glucose as osmoticum, where the rate of cell divisions under optimal culture conditions is relatively high (about 20% plating efficiency), protoplasts were drastically injured by the chilling procedure and died. In the medium with mannitol as the osmoticum instead of glucose, where the plating efficiency even under optimal conditions is rather low (about 8%), protoplasts withstand the chilling procedure. More-over, after the chilling treatment when the protoplasts were transferred to the optimal culture temperature of 29° C, the plating efficiency was raised to about 20%, which is the same level as in the glucose-containing medium without chilling. This effect was not observed when the medium in which the protoplasts were suspended during the chilling period was replaced with fresh medium. This suggests that under these conditions tomato protoplasts produce and excrete a factor in the cold that improves the vitality of the cells or stimulates cell division. The possible relationship between chilling sensitivity of tomato protoplasts and their ability to divide will be discussed.     

Continuous cultures of tomato and citron roots in vitro

Summary Initial trials with tomato-root cultures disclosed the desirability of employing a gently agitated liquid medium containing iron in the chelated form. For the normal cultivars “Ace” and “Tropic”, subcultures were best achieved by utilizing sectors that possessed one or more newly emerged laterals. Continuous cultures of a nonlateral-forming tomato mutant, “Diageotropica”, and of citron were accomplished by subculturing tips of the elongating primary roots. The tomato roots were cultured in White's medium with the Fe₂(SO₄)₃ replaced by 0.03 mM NaFeEDTA. Sustained growth of citron-root tips necessitated the use of a medium containing Murashige and Skoog salts, 7.5% sucrose, 100 mg per I each of citric acid and thiamine HCl, and 5000 mg per li-inositol. The success with citron-root cultures was extendable to all cultivars ofC. medica L., but not to otherCitrus species relatives. Both citron and “Diageotropica” root cultures manifested undiminished elongation through repeated subcultures; but neither produced laterals in response to any cultural treatments. Research was supported in part by National Science Foundation Grant OIP75-10390 and Elvenia J. Slosson Fellowship in Ornamental Horticulture.     

Inheritance of taste components in tomato

Components of tomato taste can be modeled from a set of instrumentally measured parameters. The inheritance was studied of two texture parameters with an effect of mealiness and pleasantness of taste: BFp and EBp. A recombinant inbred line population (F6) of the cross tomato cv Moneymaker x L. pimpinellifolium was analyzed for these parameters, and a molecular linkage map was developed for this population. A single QTL region on chromosome 9 was found with an effect on both BFp and EBp. The QTL explained 18.2 and 18.8% of the population variances, and 19.5 and 39.0% of the parental differences respectively for BFp and EBp.     

Genetics of actin-related sequences in tomato

Summary The genomic distribution of actin-related sequences in tomato was investigated using a cloned actin gene from soybean. Ten actin loci account for most of the hybridizing fragments observed with Southern analysis. Single loci were found on chromosomes 1, 3 and 10 and two loci on chromosome 4. One locus is linked to an unmapped isozyme marker, Sod-1. The four remaining actin loci are independent of each other and of any of the other markers tested. The number of actin loci in tomato (10) is greater than that estimated for soybean (8). As soybean is apparently a tetraploid and tomato a diploid, these results suggest that the number of actin loci has not been stable during the evolution of dicots. A number of these mapped loci lie in regions of the genome previously devoid of molecular markers and thus may be useful in basic and applied genetic research.     

Isozymic gene linkage map of the tomato: Applications in genetics and breeding

Summary New linkage data are presented for the situation of five previously unlocated isozymic loci of the tomato and closely related species with homosequential chromosomes.Prx-1 lies on chromosome 1, where it is also linked withSkdh-1; Aps-2 is linked withGot-4 on chromosome 8;Tpi-2 has been allocated to chromosome 4; and a linkage has been detected betweenPgi-1 andEst-4, whose respective chromosome has not yet been determined. These and previously published data have been summarized in the form of an isozyme linkage map. Twenty-two loci have thus been mapped on nine of the twelve tomato chromosomes. We discuss some new applications of mapped isozymic genes. In certain types of segregations, isozymic genes are far more efficient than morphological markers in providing linkage information. They greatly expedite the cytogenetic investigation of species hybrids and can be utilized to facilitate backcross transfers of genes from wild to cultivated taxa.     

Far-red light-insensitive, phytochrome A-deficient mutants of tomato

We have selected two recessive mutants of tomato with slightly longer hypocotyls than the wild type, one under low fluence rate (3 mol/m²/s) red light (R) and the other under low fluence rate blue light. These two mutants were shown to be allelic and further analysis revealed that hypocotyl growth was totally insensitive to far-red light (FR). We propose the gene symbol fri (far-red light insensitive) for this locus and have mapped it on chromosome 10. Immunochemically detectable phytochrome A polypeptide is essentially absent in the fri mutants as is the bulk spectrophotometrically detectable labile phytochrome pool in etiolated seedlings. A phytochrome B-like polypeptide is present in normal amounts and a small stable phytochrome pool can be readily detected by spectrophotometry in the fri mutants. Inhibition of hypocotyl growth by a R pulse given every 4 h is quantitatively similar in the fri mutants and wild type and the effect is to a large extent reversible if R pulses are followed immediately by a FR pulse. After 7 days in darkness, both fri mutants and the wild type become green on transfer to white light, but after 7 days in FR, the wild-type seedlings that have expanded their cotyledons lose their capacity to green in white light, while the fri mutants de-etiolate. Adult plants of the fri mutants show retarded growth and are prone to wilting, but exhibit a normal elongation response to FR given at the end of the daily photoperiod. The inhibition of seed germination by continuous FR exhibited by the wild type is normal in the fri mutants. It is proposed that these fri mutants are putative phytochrome A mutants which have normal pools of other phytochromes.     

Salt tolerance in Lycopersicon species. II. Genetic effects and a search for associated traits

Eleven quantitative traits, mostly related to tomato plant growth and fruit set, and their association with salt tolerance in terms of fruit yield under a 171.1 mM NaCl treatment have been investigated in 206 progeny derived from an interspecific hybrid, L. esculentum x L. pimpinellifolium, by self-pollination. None of the traits were highly correlated phenotypically to salt tolerance; however, the immunologically-detected presence of peptide 2 was significantly associated with high total fruit weight (TW) and number (FN) under saline treatment. Broad-sense heritability was estimated for these two salt-tolerance components as 53.44 and 72.59 %, respectively. Non-additive gene effects, which have to be considered in a breeding program for salt tolerance, have been detected in TW, FN and in average fruit weight (FW). Given that different types of gene action have been found depending on the presence or absence of a high NaCl concentration in the nutrient solution, a different set of genes, or genes, differently regulated, must be involved in the expression of TW, FN and other fruit-related characters depending on this environmental condition.