Positive correlation between menthol content andin vitro menthol tolerance inMentha arvensis L. cultivars

Menthol is a highly valued monoterpene produced by Japanese mint (Mentha arvensis) as a natural product with wide applications in cosmetics, confectionery, flavours, beverages and therapeutics. Selection of high menthol yielding genotypes is therefore the ultimate objective of all genetic improvement programmes inMentha arvensis. A positive correlation was observed in the present study between menthol content in oils of evaluated genotypes and the level of tolerance to externally supplied menthol of explants of these genotypes in culture medium. The easy use of this relationship as a selectable biochemical marker opens the practical applicability of largescalein vitro screening of the germplasm, clones and breeders' material for selection of elite genotypes.     

Effect of Sclerotinia sclerotiorum on the disease development,growth, oil yield and biochemical changes in plants of Mentha arvensis

Experiment was carried out to determine the effect of Sclerotinia sclerotiorum on the disease development, growth, oil yield and biochemical changes in the plants of Mentha arvensis. With the increase in initial inoculum levels of S. sclerotiorum a corresponding decrease in plant fresh and dry weights were recorded. The maximum reduction in the shoot-roots/suckers fresh weight and shoot-roots/suckers dry weights (39.8%, 43.6%, 40.3% and 42.9%), respectively, was observed at the highest initial inoculum level of 12 g fungal mycelium/5 kg soil as compared to uninoculated control. The infection of roots and suckers due to S. sclerotiorum increased with increasing initial inoculum levels. At the lowest initial inoculum (1.0 g mycelium/5 kg soil), infection was observed 18.0% and at the highest (12 g mycelium/5 kg soil), it was 80.2%. Significant (P ⩽ 0.01) reduction in oil yield, total chlorophyll, total phenol and total sugar content of M. arvensis plants was observed at the lowest inoculum level as compared to uninoculated control.     

Depolymerised carrageenan enhances physiological activities and menthol production in Mentha arvensis L.

Irradiated carrageenan (IC) could elicit plant growth promoting activities in plants. The effect of foliar spray of five concentrations of IC (20, 40, 60_, 80 and 100 mg L⁻¹) was studied on Mentha arvensis L. in terms of plant growth, physiological attributes, herbage yield and the content and yield of essential oil and its components. Un-irradiated carrageenan and deionized water had no effect on the attributes studied. GPC study revealed formation of low molecular weight fractions in irradiated samples containing less than 20,000 molecular weight oligomers which are responsible for plant growth promotion in this study. 80 mg L⁻¹ of IC was the most effective concentration which resulted in the highest values of growth attributes, herbage yield and the content and yield of essential oil and menthol content of the oil. It also improved the leaf-nutrient contents, photosynthetic rate and other physiological parameters. 100 mg L⁻¹ of IC did not further improve the attributes studied, but it was always better than the control.     

In vitro selection for 2,4-D tolerance in red clover

Summary In vitro, selection is a viable method of selecting herbicide-tolerant crops. This research was to evaluate in vitro selection techniques for enhancing 2,4-D [(2,4-dichlorophenoxy) acetic acid] tolerance in red clover (Trifolium pratense L.). In vivo and in vitro responses to 2,4-D of eight diverse red clover populations were correlated (r=0.77), justifying in vitro selection for 2,4-D tolerance. Suspension cultures of a red clover genotype capable of regeneration were plated onto agar-based nutrient media supplemented with 0.18 mM 2,4-D for selection experiments. After two cycles of selection, 16 2,4-D tolerant callus lines were identified based on visual growth assessment. These lines were evaluated for 2,4-D tolerance (based on 2,4-D content), using a 2,4-D bioassay procedure which consisted of placing selected callus tissue pieces on top of oat (Avena sativa L.) coleoptile or internode sections. The relative amount of 2,4-D in the callus tissue was estimated by the amount of oat section elongation after 24 h. Two of the more tolerant callus lines had 61% and 83% less 2,4-D in their tissues than the susceptible control tissue. These studies indicated that in vitro selection can enhance the levels of 2,4-D tolerance in red clover callus tissue.Florida Agricultural Experiment Station Journal Series No. 8943     

Microgametophytic selection in two alfalfa (Medicago sativa L.) clones

Summary Microgametophytic selection was investigated using two ecologically diverse autotetraploid clones of alfalfa. Several selection pressures (drying, aging, freezing, and high and low temperatures) were applied to microgametophytes at three stages of the life cycle, 1) during microsporogenesis, 2) post-anthesis, and 3) pollen tube growth. Pollen aging produced a progeny population with a greater mean plant size and a lower coefficient of variation than the control progeny. High temperature (29.5 °C) applied both during microsporogenesis and pollen tube growth resulted in progeny populations which were significantly taller and, in one case, had a larger leaf number than the control populations. In contrast, air dried pollen resulted in a progeny population which had significantly smaller character means and larger coefficients of variation than the control population. Also, low temperature (15 °C) during pollen tube growth yielded progeny with reduced branch number and a larger coefficient of variation than the control progeny. In cases where progeny derived from selected microgametophytes were found to differ from the control offspring, corresponding shifts in the reciprocal cross were not observed. For the temperature stress treatments, the lack of reciprocal differences may be related to the different temperature adaptations of the two ecotypes. These results suggest that microgametophytic selection can be effective in shifting the mean of the progeny generation; however, the results obtained will vary depending upon the selection pressure, stage of selection, and the parents used.     

Effect of nitrogen and phosphorus status on the translocation of three herbicides in field bindweed (Convolvulus arvensis L.)

Twenty-eight day old field bindweed plants grown in culture solutions deficient in nitrogen (N) or phosphorus (P) for the last seven days of growth translocated significantly less foliarly applied dicamba (3,6-dichloro-o-anisic acid) and 2,4-D [(2,4-dichlorophenoxy) acetic acid] to their roots than did plants grown in complete nutrient solutions. In contrast, N deficiency stimulated basipetal translocation of glyphosate [N-(phosphonomethyl) glycine] and inhibited its acropetal translocation in field bindweed. Deficiencies of both N and P decreased translocation of dicamba from the treated area, but had no influence on translocation of glyphosate or 2,4-D from the treated area.Journal Article No. 4406 of the Agric. Exp. Stn., Oklahoma State University.     

Stimulation of menthol production in <Emphasis Type="Italic">Mentha piperita</Emphasis> cell culture

Plant cell culture provides an alternative means for producing secondary metabolites. In this study, experiments were carried out to study the impact of several parameters, independently and in combination, on the stimulation of menthol production in the cell suspension culture of Mentha piperita. Callus was obtained from leaf segments of in vitro grown plantlets on Murashige and Skoog (MS) medium supplemented with 0.2 mg l⁻¹of 2,4-dichlorophenoxy acetic acid to initiate cell suspension culture. This culture was maintained in half-strength MS medium supplemented with 0.2 mg l⁻¹of 2,4-dichlorophenoxy acetic acid at 15 d interval and used for further studies. Precursor feeding alone, i.e., menthone, at 35 μM concentration showed slightly improved productivity. γ-Cyclodextrin alone at 60 μM concentration and in combination with menthone feeding at 35 μM increased menthol yield up to 92 and 110 mg l⁻¹ in comparison to 77 mg l⁻¹ of control culture. Synergistic potentiation effect of menthone feeding at 35 μM and γ-cyclodextrin at 60 μM treatment followed by in situ adsorption with RP-8 also showed potential stimulation of menthol production in M. piperita cell culture. Fungal elicitor treatment showed enhanced production level up to 140.8 mg l⁻¹ in comparison to that of control. Further studies were carried out with the establishment of Agrobacterium tumefaciens (Ach5) gall-mediated calli, and consequently, cell suspension culture and results showed the significant enhancement of menthol yield up to 278 mg l⁻¹. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

In vitro selection of a glyphosate-tolerant sugarcane cellular line

Cell suspensions derived fromSaccharum spp. varieties (PR62258, V64-10, V71-51) were used to study the effects of glyphosate on cell suspensions of commercial cultivars of sugarcane in order to select and develop a glyphosate-tolerant cellular line. A liquid medium with 0.8 mM/L glyphosate was used for selection. The results showed growth inhibition of approximately 50% for cell suspensions in mediums with 0.8 mM/L glyphosate. The results obtained in vitro were consistent with the results reported from in vivo tests. Plants were regenerated from sensitive cell suspensions and tolerant cell suspensions from cultivar V71-51. Tolerant cell suspensions tolerated a concentration that was 12.5-fold higher than that of the sensitive cell suspensions. Plants regenerated from tolerant cell suspensions tolerated a glyphosate concentration that was 6-fold higher than that of the plants regenerated from sensitive cell suspensions. Cell suspensions derived from tolerant regenerated plants tolerated a glyphosate concentration that was 5-fold higher than that of those derived from sensitive regenerated plants. The RAPD patterns obtained with OPA-07 revealed a 564-bp band that can be used to characterize the tolerant cellular line. This band is not present in the sensitive cellular line. Consequently, the tolerance persisted throughout the differentiation into plants and dedifferentiation into cell suspensions. This performance suggests that an in vitro selection of pre-existing variability has taken place.     

In vitro characterization of salt stress effects and the selection of salt tolerant plants in rice (Oryza sativa L.)

Summary The response of plant cells to salt stress was studied on embryo derived calli of rice (Oryza sativa L.) in order to identify cellular phenotypes associated with the stress. The feasability of selecting salt tolerant callus and its subsequent regeneration to plants was also studied. Callus was grown on agar-solidified media containing 0%, 1% and 2% (w/v) NaCl for 24 days. Parameters such as fresh weight, dry weight, soluble protein and proline content were measured. The callus growth decreased markedly with increasing NaCl concentration in the medium. The proline content was enhanced several fold in salt stressed calli. A prolonged exposure of callus to the salt environment led to discolouration and arrested growth in the majority of the calli and only a small number of callus cells maintained healthy and stable growth. These variants were subcultured every three weeks for a period of four months onto medium containing 1% NaCl to identify tolerant lines. At the end of the third cell passage, the tolerant calli were transferred to regeneration medium to regenerate plants. The regeneration frequency in the salt-selected lines was enhanced when compared to unselected lines.     

Sporophytic response to pollen selection for Alachlor tolerance in maize

In order to assess the efficiency of male gametophytic selection (MGS) for crop improvement, pollen selection for tolerance to herbicide was applied in maize. The experiment was designed to test the parallel reactivity to Alachlor of pollen and plants grown in controlled conditions or in the field, the response to pollen selection in the sporophytic progeny, the response to a second cycle of MGS, and the transmission of the selected trait to the following generations. The results demonstrated that pollen assay can be used to predict Alachlor tolerance under field conditions and to monitor the response to selection. A positive response to selection applied to pollen in the sporophytic progeny was obtained in diverse genetic backgrounds, indicating that the technique can be generally included in standard breeding programs; the analysis of the data produced in a second selection cycle indicated that the selected trait is maintained in the next generation.     

An in vitro whole plant selection system: paraquat tolerant mutants in the fern Ceratopteris

Summary A whole plant selection system using the haploid gametophyte generation of the fern Ceratopteris richardii has been developed to select for mutations that confer resistance or tolerance to various selection pressures. The expression of the mutations can be analyzed and characterized in both the haploid gametophyte and diploid sporophyte generations. Genetic analyses are facilitated by the fern's rapid life cycle and the ease of manipulating the gametophyte generation. Selection for tolerance to the herbicide paraquat has yielded two mutants which have an increased tolerance to the herbicide in both the gametophyte and sporophyte generations. Both mutants exhibit single nuclear gene inheritance patterns and appear to be closely linked or allelic.     

Anticytokinin effects on in vitro response of embryogenic and nonembryogenic genotypes of Dactylis glomerata L.

The effect of triazine and carbamate type of anticytokinins on in vitro response of embryogenic and nonembryogenic genotypes of orchardgrass (Dactylis glomerata L.) was studied. Both compounds stimulated callus production. Anticytokinins influenced both the efficiency of somatic embryogenesis and frequency of embryoid formation.     

Agronomical and chemical characterization of spearmint (Mentha spicata L.) originating in Turkey

The essential oil properties of spearmint (Mentha spicataL.), one of the most important spice plants, were studied and the essential oil components determined using gas chromatography. The essential oil content of wild-grown spearmint in the region was found to range from 1.00% to 2.00%, and two chemotypes were identified, one high in carvone (49.53-80.65%) and the other in pulegone (44.9-49.23%). Agronomic and essential oil properties of cultivated landraces ofM. spicata were also investigated under field conditions during the 1999 vegetation period. The examined spearmint landraces showed a great variability for each character studied, including yield and essential oil components. The crop was harvested twice during the vegetation period, and the essential oil content of the landraces varied from 0.90 to 2.70% in the first harvest and from 1.00 to 3.00% in the second one. Carvone was constantly present as the predominant essential oil in landraces, except for one sample, which was high in linalool (82.80%). Superior landraces with carvone contents were discovered; their maximum content reached 79.70% in the first cutting and 82.97% at the second cutting. The superior landraces were assayed for future improvement studies.     

A major locus for submergence tolerance mapped on rice chromosome 9

Submergence stress is a widespread problem in rice-growing environments where drainage is impeded. A few cultivars can tolerate more than 10 days of submergence, but the genes conferring this tolerance have not been identified. We used randon-amplified polymorphic DNA (RAPD) and restriction fragment length polymorphism (RFLP) markers to map submergence tolerance in 169 F₂ plants and the resulting F₃ families of a cross between a tolerant indica rice line, IR40931-26, and a susceptible japonica line, PI543851. IR40931-26 inherited strong submergence tolerance from the unimproved cultivar FR13A. Eight-day old F₃ seedlings were submerged for 14–16 days in 55-cm deep tanks, and tolerance was scored after 7 days recovery on a scale of 1 (tolerant) to 9 (susceptible). The tolerant and susceptible parents scored 1.5 and 8.4, respectively, and the F₃ means ranged from 1.6 to 8.9. Two bulks were formed with DNA from F₂ plants corresponding to the nine most tolerant and the nine most susceptible F₃ families. Of 624 RAPD primers used to screen the bulks, five produced bands associated with either tolerance or susceptibility. These markers were mapped to a region of chromosome 9 by linkage to RFLP markers. A submergence tolerance quantitative trait locus (QTL), here designatedSub1, was located ca. 4 cM from the RFLP marker C1232 and accounted for 69% of the phenotypic variance for the trait.     

Characterization of progeny of Coleus blumei following an in vitro selection for salt tolerance

An in vitro selection method was developed for Coleus blumei to enhance salt tolerance of this amenity species. Leaf disc explants were incubated on a Murashige & Skoog medium containing benzylaminopurine, 2 mg l^-1, and napthalene acetic acid, 1 mg l^-1, which initiated both callus and plantlets from the explants. A large number of explants were incubated on this differentiating medium containing 90 mM NaCl, which inhibited over 90% of plantlet formation. Surviving plantlets. were grown to maturity, when apical cuttings were taken and propagated. Plants were also allowed to flower and set seed. Cuttings from the selected regenerated plants showed consistently better growth in the presence of NaCl than unselected cuttings. Seed progeny of selected plants also showed more vigorous growth in the presence and absence of NaCl than progeny from unselected plants. The in vitro selection was compared with the results of an earlier in vivo selection to assess the contribution from tissue culture derived somaclonal variation. Progeny from the in vitro selection showed a higher level of tolerance than progeny from the in vivo selection.     

A new approach to consider the pollen variable in forecasting yield models

Methods for forecasting harvest yields have been improved considerably in the last 20 years with the development of new data survey (remote sensing) and statistical techniques. One of these methods, based on pollen release in the atmosphere, is especially important for anemophilous species such as olive. The aim of the present work is to use a different approach to forecast the olive harvest by considering the pollen variable as “endogenous” because it is involved in the consequential processes from the formation of pollen to fruiting, the complex of which determines, more or less, the final production. Unlike models built upon a single equation (multiple linear regression analysis), the proposed estimate, based on an incomplete system of equations, recovers the consistency associated with the inference of parameters while avoiding the errors of “over-estimation.” The study, based on 17 years of data considers the quantity of olive pollen monitored and the relative annual olive production in addition to climatic, agronomic, and pathological variables associated with production. The harvest forecast provides the possibility for planning and optimizing the various stages of olive production from cultivation to distribution, including sound management of the olive supply.

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Une nouvelle approche pour considérer la variable pollen dans les modèles de prévision des rendement de récolte

Résumé  Pendant les vingt dernières années les méthodes de prévision des rendements de récolte ont été considérablement améliorées grace au développement de nouvelles techniques statistiques et d’ enquête des données (télédétection). Parmi ces méthodes celle basée sur l’émission du pollen dans l’atmosphère se révèle particulièrement importante pour les espèces anémophiles comme l’olivier. Le but de ce travail est celui d’arriver à employer une méthode différente dans la prévision des rendements de récolte de l’olivier, le tout en considérant comme “endogène” la variable pollen. Ce dernier est, en effect, impliqué dans les processus d’évolution qui vont de sa formation à la fructification, de manière à déterminer la production finale. Contrairement aux modèles établis sur une équation simple (analyse multiple de régression linéaire), l’évaluation proposée, basée sur un système inachevé d’équations, récupère la consistance connexe à l’ inférence des paramètres tout en évitant les erreurs de “surestimation”. L’étude, basée sur dix-sept ans de données, considère la quantité de pollen d’olivier détectée et conséquemment la production oléicole annuelle, outre aux variables climatiques, agronomiques et pathologiques liées à la production. Les modèles de prévision offrent la possibilité de rationaliser les différentes phases de la filière oléicole en optimisant les procédés, de la production à la distribution, y compris la gestion rationnelle des stocks.

     

Autofertility and rate of cross-fertilization: crucial characters for breeding synthetic varieties in faba beans (Vicia faba L.)

Summary To increase the level and stability of yield in faba beans (Vicia faba L.), heterosis must be exploited. Hybrids are not available because of the instability of male sterility. Synthetic varieties can and should be bred. Thus, we studied the reproductive behavior of this partially allogamous, insect-pollinated crop. Autofertility (AF) and the rate of cross-fertilization (C) were measured in 36 inbred lines and 28 crosses in F₁, F₂, and F₃ generations for 3 years at Hohenheim, Stuttgart, FRG. Heterozygosity strongly increased AF and decreased C. AF was negatively correlated with C. AF varied from 1% to 98%, and C varied from 7% to 82%. Heritability for both characters was high. For an optimum exploitation of heterosis, breeders should utilize lines with high C as variety components. It is often labor-intensive to multiply such lines, due to low AF. Hence, breeders tend to use autofertile lines with rather limited C. We showed that even in this case about 50% of the total heterosis, which equals a yield increase of at least 25% over the inbred line level, is realized. An increase in yield stability due to heterogeneity will occur simultaneously.     

Genetic studies of self-fertility in rye (Secale cereale L.). 1. The identification of genotypes of self-fertile lines for the Sf alleles of self-incompatibility genes

Segregation for self-fertility has been studied in progenies from the crosses of self-sterile (SS) plants with interline hybrids obtained by a diallel scheme of pollinations between seven self-fertile (SF) lines (nos. 2–8) and with F₁ (SS plant x SF line) hybrids. All the offspring families from the SS plant x F₁ (SS plant x SF line) crosses demonstrated a 1SF1SS segregation. The crosses of SS plants with some interline hybrids gave only self-fertile plants, whereas the crosses with other interline hybrids gave a segregation of 3SF:1SS expected in the case of digenic segregation. The data obtained permitted us to identify three different S loci (S1, S2, S5) and to estimate the genotypes of self-fertile lines for their Sf alleles: lines 5, 6, 7 and 8 are S1f/S1f S2n/S2n S5m/S5m, line 4 is S1n/S1n S2f/S2f S5m/S5m, and lines 2 and 3 are S1n/S1n S2m/S2m S5f/S5f(Sn, Sm designate active alleles of the incompatibility genes). The identification of the particular S gene which is presented by the Sf allele in each line has been made on the basis of our data concerning the linkage of the Sf mutation with isozyme markers of particular rye chromosomes, which is reported in an accompanying paper.     

Rapid evaluation of sorghum for aluminum tolerance

Two screening methods tested in this study were based on the observation that roots of freshly germinated seeds exhibit their relative tolerance to Al during the first 48 hours of growth. Sorghum (Sorghum bicolor L. Moench) varieties were evaluated using toxic/nontoxic soil pairs in petri dishes and toxic/nontoxic solution pairs in a flowing aqueous system. The soils had 0, 61, and 72% Al saturation and the solutions had 0, 1.85 and 3.70 M l^–1 Al from AlCl₃ and 0.25 mM l^–1 Ca from Ca(NO₃)₂. Relative root lengths in both systems correctly differentiated 13 genotypes of known Al tolerance into tolerant and intolerant groups. Twenty six other sorghums genotypes were also screened using genotypes of known Al reactions as checks. The soil with 72% Al saturation and the 1.85 M l^–1 Al solution gave the highest correlation between the two systems and both effectively arrayed sorghums of known and unknown tolerance. For routine screening the authors prefer the soil system for its simplicity, efficiency, and use of a natural growth medium.Journal paper 11637. Purdue University Agricultural Experiment Station, Lafayette, Indiana. Contribution from the Agronomy Department.     

In vitro system for studying the interaction between Erwinia amylovora and genotypes of pear

A new in vitro system is described for studying an interaction between Erwinia amylovoraand Pyrus communis (L.). The system uses single shoots placed onto the solid medium, and it enables to detect changes in pH of the medium and differential appearance of shoot necrosis. Shoots of susceptible cultivar (Williams) and tolerant cultivar (Harrow Sweet) were compared measuring the necrosis rate along the in vitroshoots and the pH variation following proton extrusion of both plant and pathogen. Shoots acidified differentially the culture medium depending on the presence of the pathogen, cultivar susceptibility and shoot inoculation methods. Differences in the tolerance level against pathogen among the cultivars were distinguishable only when the shoots were inoculated at the basal end. In susceptible cultivar, the necrosis appeared after 48 h of inoculation, while in tolerant cultivars after 72 h. This system is repeatable and more reliable than already known methods, such as in vitroleaf explants or in vivoplants; it can be used all around the year to test the gene expression and products essential to characterize the genes involved in the pathogenesis. This system showed the effects of E. amylovoraon the photosystem dependent system of host cells, confirmed by the effects of pathogen attack on the variation of chlorophyll a and chlorophyll b ratios and positive effects of light on the appearance of the first disease symptoms.