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.     

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     

Genetic variability in Lycopersicon species and their genetic relationships

Summary Genetic diversity has to be described and measured in order to establish breeding strategies and manage genetic resources. It is also fundamental to develop a comparative intraspecific study before attempting to discuss and conclude any phylogenetic relationship. The genetic variability of Lycopersicon species was studied using starch gel electrophoresis of 11 enzymatic systems in a hierarchical fashion. The species with the greatest genetic variability are L. chilense, L. peruvianum and L. pennellii, mainly due to the within-line component. L. chmielewskii, L. parviflorum and L. pimpinellifolium show an intermediate total variability and their between-component clearly predominates over the within-component. The least variable species are L. cheesmanii and L. esculentum. Cluster analysis resulted in three main groups: one formed by the cultigen, L. pimpinellifolium, L. cheesmanii and L. peruvianum;another by two species with self-incompatibility systems, L. pennelli and L. chilense; and another by two autogamous species L. chmielewskii and L. parviflorum. With respect to L. esculentum the farthest related species is Solanum rickii and the closest, L. pimpinellifolium.     

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.     

Salt tolerance in Lycopersicon species. I. Character definition and changes in gene expression

Salt tolerance defined in terms of fruit yield under different NaCl concentrations (171.1 and 325.1 mM) is analyzed in 11 lines belonging to: Lycopersicon esculentum, L. cheesmanii, L. chmielewski, L. peruvianum and L. pimpinellifolium. Four L. pimpinellifolium lines and two L. cheesmanii lines tolerated the 171.1mM treatment; the latter species even tolerates 325.1 mM of NaCl. Changes in gene expression induced by salt treatment were also investigated by studying anther and leaf zymograms for L. esculentum and one salt-tolerant L. pimpinellifolium line, and leaf proteinograms for all lines. Changes in leaf PRX and MDH enzymatic systems were detected, mainly in the salt-sensitive genotype (L. esculentum). Four saltrelated peptides from 14 500 to 40 000 daltons were found. A polyclonal antibody raised against one of these peptides (number 2), also binds another peptide, named 2, of much higher molecular weight, present both in control and salt-tolerant L. cheesmanii lines at the end of 171.1 mM treatment. The xero-halophyte shrub Atriplex halimus also showed a likely 2-homologous peptide with this treatment, while its counterpart C₃ species A. triangularis did not.     

Genetics of esterases in Drosophila. IV. Slow-migrating S-esterase in Drosophila of the virilis group

A slow-migrating -esterase (S-esterase) is described which has been detected in Drosophila montana, Drosophila imeretensis, and some stocks of Drosophila virilis when mixtures of - and -naphthyl acetate are used as substrates in histochemical reactions after electrophoresis. Sexual dimorphism for S-esterase has been demonstrated. This esterase is contained in male genitalia only, predominantly in the ejaculatory bulb (waxy plug). It appears 3–4 days after emergence of flies. In hybrids between S⁺ and S⁰ species, the activity of the slow esterase is either decreased or inhibited. An autonomous synthesis of the S-esterase in the ejaculatory bulb was established by transplantation of imaginal genital discs into larvae of different Drosophila stocks. Based on analysis of physicochemical and immunochemical properties, S-esterase is suggested to be an independent fraction of esterase, possibly dimeric, which does not cross-react with -esterase antiserum.     

Extent of genetic variability of endosperm esterases in Triticum aestivum L. 2n=6x=42

Summary Genetic variability of endosperm esterase has been studied in 42 cultivars of Triticum aestivum L. 2n=6x=42. Different techniques, including sequential electrophoresis and electrofocusing, have been used with various substrates and esterase inhibitors. The electrophoretic patterns in each cultivar are described. Chromosomal location using the nullitetrasomic and ditelosomic lines of Chinese Spring was carried out in order to relate and/or locate the esterase genes to specific chromosomes. Most of the esterase isozymes located were in the long arm of the chromosomes of the homoeology group 3; but we have found six located in the short arms, five of them in the chromosome 3AS and one in the 3DS. This location increases the number of esterase genes described, because no esterase genes had been described so far in short arms of chromosomes of the homoeology group 3. The genetic control is discussed and, according to our results, between 12 and 15 loci, organized in five compound loci, control the endosperm esterases in wheat. Also one modifier gene modifying the mobility of two esterase bands and present in all the cultivars studied is postulated.This work was supported by a personal grant (L. Rebordinos) from the P.F.P.I. and by an institutional grant from the C.A.I.C.Y.T. (PB85-0153)     

Somatic embryogenesis from Lycopersicon peruvianum leaf mesophyll protoplasts

Summary One to five percent of Lycopersicon peruvianum (L.) Mill. leaf mesophyll protoplasts undergo cell division and concomitant organization to form embryogenic-like structures when cultured in Murashige and Skoog medium (1962) containing 3% sucrose, 9% mannitol, 1.0 mg/l kinetin (K) and 1.0 mg/l naphthalene acetic acid (NAA) at pH 5.6–5.8 (medium A). These embryogenic structures, after passing through developmental stages similar to those observed in zygotic embryogeny, are capable of forming shoots on hormone-free medium A. In medium B, wherein 0.5 mg/l of 2,4-dichlorophenoxyacetic (2,4-D) replaced the hormones (K and NAA), embryogenic structures did not develop. However, callus originating in medium B retained morphogenetic capacity as was evidenced by subsequent shoot regeneration when they were transferred to medium A with K and NAA replaced by 1.0 mg/l zeatin (Z). The potential value of incorporating this regeneration trait into Lycopersicon species and cultivated lines for use in tissue culture programs is discussed.Michigan Agricultural Experiment Station Journal No. 9676     

Salt tolerance in Lycopersicon species. III. Detection of quantitative trait loci by means of molecular markers

A segregating population derived from a cross between L. esculentum cv Madrigal and a line of L. pimpinellifolium was used to identify genetic markers linked to QTLs involved in salinity tolerance in terms of yield, under a conductivity of 15 dS/m (171.1 mM NaCl). Six markers resulted, associated with QTLs affecting average fruit weight, fruit number and total weight under salinity. One of them, Aco-1, behaves reversely to the expectation from parental means; this and other features make it a promising target to obtain salt-tolerant tomatoes. Epistatic interactions were also found, thus affecting the criteria for marker-assisted selection. Although only 41% of the loci assayed were polymorphic, a high efficiency in identifying QTLs was achieved, since 43% of the marker loci are linked to QTLs for the trait under study.     

Screening of wild accessions resistant to gray mold (Botrytis cinerea Pers.) in Lycopersicon

Tomato gray mold (Botrytis cinerea Pers.) is a common disease worldwide, and often causes serious production loss by infecting leaves, stems, flowers and fruits. Presently, no resistant cultivars are available. To find new breeding materials for gray mold resistance, assessment for resistance of the leaflet and stem in six tomato cultivars, 44 wild tomato accessions and a Solanum lycopersicoides accession was performed. Although no correlation was observed (r=−0.127^ns) between resistance of the leaflet and the stem, L. peruvianum LA2745, L. hirsutum LA2314 and L. pimpinellifolium LA1246 showed high resistance both in the leaflet and in the stem. Particularly, in the leaves of LA2745, no lesions were observed even more than two weeks after the inoculation with conidia, and F1s between a cultivated tomato and LA2745 also showed high resistance as observed in LA2745. From these results, LA2745 is thought to be a promising material for breeding gray-mold resistant cultivars.     

Evolution of fraction 1 protein in the genus Lycopersicon

The large- and small-subunit polypeptide composition of fraction 1 protein contained in seven species of Lycopersicon and Solanum pennellii was determined by electrofocusing. The eight species of protein had large subunits composed of three polypeptides separated by about 0.05 pH unit, but there was no difference in the isoelectric points of the clusters of three polypeptides. By this criterion, no surviving mutations have appeared in the extranuclear DNA coding for the cluster of large-subunit polypeptides during a period of evolution which generated the eight species of plants. The genus Lycopersicon appears to be much younger than its sister genus Nicotiana in the family Solanaceae, where four types of polypeptide clusters have evolved. Three different small-subunit polypeptides whose isoelectric points are coded by nuclear DNA have arisen among the seven Lycopersicon species, and L. hirsutum and S. pennellii have proteins containing single polypeptides and are therefore considered older than L. chilense, L. chimielewskii, and L. parviflorum, whose proteins contain two polypeptides. L. cheesemanii, L. pimpinellifolium, and L. esculentum (and probably L. peruvianum) seem to be the most recently evolved species since their fraction 1 proteins have small subunits composed of three polypeptides.This research was supported by NSF Grant 75-07368 and Contract No. EY-76-S-03-0034, P. A. #8, from the Department of Energy.     

Sources and inheritance of resistance to leaf curl virus in Lycopersicon

Summary One hundred and twenty-two varieties, lines and wild accessions of Lycopersicon were screened under three different regimes during the autumn/winter season of 1982–83 and 1983–84 for resistance to tomato leaf curl virus (TLCV). L. hirsutum f. glabratum (B6013) and L. hirsutum f. typicum (A1904) proved to be highly resistant to TLCV in all three environments. Various accessions of L. peruvianum were also highly resistant. L. pimpinellifolium (A1921) exhibited no TLCV symptoms within 90 days. Of the cultivated varieties, Acc 99 exhibited the minimim score for susceptibility; AC 142, Collection No. 2, Kalyanpur Angurlata and HS 101 had a low rating for virus incidence. The inheritance of resistance was studied in the interspecific crosses between a TLCV resistant line of L. pimpinellifolium (A1921) and five (HS 101, HS 102, HS 110, Pusa Ruby and Punjab Chhuhara) susceptible cultivars of L. esculentum. Parents, F₁, F₂ and backcross progenies were artificially inoculated with local strains of TLCV using vector the viruliferious whitefly, Bemisia tabaci (Genn.). Data indicated that the resistance of L. pimpinellifolium (A 1921) is monogenic and incompletely dominant over susceptibility.     

Analysis of the organogenetic potential of calli of three Canary Island Lycopersicon esculentum land races

The organogenetic potential from callus of three tomato land races from the Canary Islands adapted to semi-arid environment (Salvaje, Rusa and Especial), and one tomato cultivar (Meltine), were examined. The response of four explant types (cotyledon, shoot apex, hypocotyl and root) to nine PGR regimes (BAP at 1 or 2 or 5 mg/l) + either IAA (0.5 mg/l) or 2,4-D (0.5 or 1 mg/l) were measured. BAP at 5 mg/l+IAA at 0.5 mg/l induced most organogenesis in all the explant types for all genotypes. Salvaje has one of the highest organogenetic potentials described in tomato.Abbreviation OP = organogenetic potential     

RFLP analysis of phylogenetic relationships and genetic variation in the genus Lycopersicon

Summary Forty single-copy, nuclear probes of known chromosomal position were used to examine restriction fragment length polymorphism in the tomato genus Lycopersion. The probes were from three libraries: one cDNA, and two genomic libraries ne genomic made with EcoRI and the other with PstI. Total DNA from 156 plants representing eight species was cut with five different restriction enzymes and scored in 198 probe-enzyme combinations. Genetic distances between accessions (populations) and species were calculated from the resultant restriction patterns and proportion of shared bands. Accessions belonging to the same species largely clustered together, confirming their current classification. However, one mountain accession, classified as L. peruvianum var. humifusum (LA2150), was sufficiently distinct from the other accessions of L. peruvianum that it may qualify as a separate species L. esculentum and L. pimpinellifolium were the least clearly differentiated, possibly reflecting introgressive hybridization, known to have been promoted by man in recent history. Dendrograms constructed from cDNA versus genomic clones were nearly identical in their general grouping of species. The dendrograms revealed two major dichotomies in the genus: one corresponding to mating behavior [self-compatible (SC) versus self-incompatible (SI) species] and the other corresponding to fruit color (red versus green-fruited species). The ratio of withinversus between-accession diversity was much lower for SC species, indicating that most of the diversity within these species exists between populations, rather than within populations. Overall, the amount of genetic variation in the SI species far exceeded that found in SC species. This result is exemplified by the fact that more genetic variation could be found within a single accession of one of the SI species (e.g., L. peruvianum) than among all accessions tested of any one of the SC species (e.g., L. esculentum or L. pimpinellifolium). Results from this study are discussed in relationship to germ plasm collection/utilization and with regard to the use of RFLPs in tomato breeding and genetics.     

Identification and distribution of profilin in tomato (Lycopersicon esculentum Mill.)

Profilin is a G-actin monomer-binding protein which has been shown to participate in actin-based tipgrowth of animal cells. The abundance of profilin in pollen and its occurrence in several vegetable foods raises the question of the role of profilin in plants. First, its distribution throughout various parts of the plant needs to be determined. This paper describes observations on the presence of profilin in the tomato plant (Lycopersicon esculentum Mill.). The distribution of profilin in flower buds, stems, leaves, roots, and fruits of tomato was determined by immunoblotting and by tissue printing, showing that profilin is present in most if not all parts of the tomato plant.We gratefully acknowledge the help provided by Dr. A.T. Jagendorf and the donation of tomato seeds and maize pollen by N. Eanetta and Dr. M. Smith, respectively. The use of Dr. R. Wayne's SZH ILLD dissecting microscope is gratefully acknowledged. This work was aided by helpful discussions with C.S. Combs, Dr. C.A. Conley, and Dr. J. Andersland. This work was supported by a Hatch grant and NRI Competitive Grants Program/USDA 94-37304-1046 to MVP. This material is based upon work supported under a National Science Foundation Graduate Research Fellowship to DWD.     

Compatibility studies in three wild species of Lycopersicon

A study was made on the self-compatibility in L. hirsutum (LH), L. pennellii (LP) and L. chilense (LC) and on cross-compatibility or preferable unilateral compatibility between these wild species and L. esculentum (LE). From the results of various test crosses and selfings and of cytological research and fruit and seed setting as well it was concluded that the material used is compatible but it differed in the degree of the expression. The highest self- and cross-compatibility was found in LP where number of seed, their viability and also pollen tube growth was similar to LE. Differences between LC and LH in self-compatibility and the congruity with LE were very small on the basis of pollen tube elongation but relatively fewer number of fruit and seed were obtained in the cross LE × LC than from the combination of LE × LH. In the reciprocal crosses when wild species were used as the pistillate parent, no seed were obtained as fertilization was prevented.     

Extremely thermostable esterases from the thermoacidophilic euryarchaeon Picrophilus torridus

Two genes encoding esterases EstA and EstB of Picrophilus torridus were identified by the means of genome analysis and were subsequently cloned in Escherichia coli. PTO 0988, which is encoding EstA, consists of 579 bp, whereas PTO 1141, encoding EstB, is composed of 696 bp, corresponding to 192 aa and 231 aa, respectively. Sequence comparison revealed that both biocatalysts have low sequence identities (14 and 16%) compared to previously characterized enzymes. Detailed analysis suggests that EstA and EstB are the first esterases from thermoacidophiles not classified as members of the HSL family. Furthermore, the subunits with an apparent molecular mass of 22 and 27 kDa of the homotrimeric EstA and EstB, respectively, represent the smallest esterase subunits from thermophilic microorganisms reported to date. The recombinant esterases were purified by Ni²⁺ affinity chromatography, and the activity of the purified esterases was measured over a wide pH (pH 4.5–8.5) and temperature range (10–90°C). Highest activity of the esterases was measured at 70°C (EstA) and 55°C (EstB) with short pNP-esters as preferred substrates. In addition, esters of the non-steroidal anti-inflammatory drugs naproxen, ketoprofen, and ibuprofen are hydrolyzed by both EstA and EstB. Extreme thermostability was measured for both enzymes at temperatures as high as 90°C. The determined half-life (t _1/2) at 90°C was 21 and 10 h for EstA and EstB, respectively. Remarkable preservation of esterase activity in the presence of detergents, urea, and commonly used organic solvents complete the exceptional phenotype of EstA and EstB.     

Genetics of the hemolymph esterases ofLucilia cuprina (Diptera: Calliphoridae)

When hemolymph from adults ofLucilia cuprina was partitioned on native polyacrylamide gels, nonspecific esterase staining demonstrated 10 bands with up to six bands in an individual. The bands derive from alleles at two loci,E _HA (five alleles) andE _HB (four alleles).E _HA is located on chromosome 4, 16.3 map units fromsv (singed vibrissae) and 22.1 map units fromra (radial vein gaps).E _HB is located on chromosome 5, 34.0 map units fromto ² (topaz² eyes) and 7.2 map units frommv (M1-veinless).