Callus and shoot formation in organ and tissue cultures of Hedera helix L., English ivy

When shoots of young plants of hemp (Cannabis sativa L.) and spinach (Spinacea oleracea L.) were cultured as cuttings and allowed to regenerate advenitious roots, ca. 80–85% became female (formed pistillate flowers) regardless of whether the leaves were left on the plants or were cut off (except for the 2–3 uppermost ones) after the beginning of adventitious-root formation. But when the leaves were cut off and the cuttings treated with gibberellic acid (GA₃, 25 mg/l) ca. 77–80% of the plants became male (formed staminate flowers). The result was quite similar when roots and leaves of young hemp plants were removed at the same time and the cuttings treated with GA₃. It is suggested that the leaves play an essential role in sex expression in hemp and spinach and that this role is related to gibberellin synthesis in the leaves.     

Aminooxyacetic acid inhibits antheridiogenesis and development of Anemia phyllitidis gametophytes

Cytomorphological studies of the development of young fern gametophytes (Anemia phyllitidis) have been used to investigate combined effects of gibberellic acid and ethylene on male sex expression. ACC (the key by-product in ethylene biosynthesis pathway) was found to exert a synergetic effect on the gibberellic acid-induced antheridia formation, and this phenomenon could be related with the specific stimulation of cell growth and activity of their differentiation. To complete and verify those observations male sex expression in the fern gametophytes treated with ACC-biosynthesis inhibitor was reinvestigated. Aminooxyacetic acid (AOA) restrained antheridia formation via inhibition of cell divisions. AOA influenced the arrangement and flexibility of cellulose microfibrils in the antheridial zone cells, thus affecting cell expansion. On the other hand, the level of DNA synthesis was not reduced. Transient increase in the number of S-phase cells, followed by the accumulation of G2-phase cells led to the enhancement of cell polyploidization. All these findings correspond with the previous observations and support participation of ethylene in gibberellic acid-induced male sex expression in ferns.Abbreviations AOA Aminooxyacetic acid - CPA Cell profile area - GA Gibberellin - GA₃ Gibberellic acid     

On the genetic control of the regeneration response in the dioecious Melandrium album

Summary Melandrium album (2n = 24), a dioecious species with heteromorphic sex chromosomes (XY, male; XX, female), has a strong genetic commitment for sex determination. So far the in vitro regeneration response in this species has appeared to be restricted to female plants. We report here a procedure for achieving active regeneration from explants and mesophyll protoplasts of selected female plants. Shoots were frequently induced in hormone-free media and in the presence of cytokinins or low concentrations (<0.5 mg/l) of auxins. The in vitro conditions employed did not affect the sex expression of the regenerated plants. Crosses between selected female plants with high regeneration capacity and nonregenerating males indicated that the regeneration response is genetically controlled and can be transmitted sexually into male and female progenies.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - 2iP (2-isopentenyl)-adenine - IAA indole acetic acid - IBA indole butyric acid - NAA naphthalene acetic acid - BAP 6-benzylaminopurine     

Study of the inheritance of sex expression in the cucumber. The interaction of major genes with modifying genetic and non-genetic factors

Summary Two major genes and a complex of polygenes affecting sex expression inCucumis sativus L. as well as their interaction with some nongenetic factors were investigated in the present study. The genetic factorst was found to affect sex by inducing a shift of the predetermined flowering pattern of this plant, in the direction of its base. As this pattern is composed of a staminate stage followed by a mixed (staminate-pistillate) stage and a pistillate stage, a double dose ofst will induce a change from the normal monoecious sex expression (to absolute gynoecism.The second major gene studied,m, known previously to control sex in the individual flower (m/m—andromonoecious,M—monoecious), also interacts with factors affecting the flowering pattern by inducing male tendency.Using an ordinary commercial monoecious stock additional modifying factors for sex expression were demonstrated, by eight generations of selection for high (in male direction) and for low (in female direction) node number to the first pistillate flower. A comparison of the means and frequency distributions of node number of the two selected lines, their F₁ and F₂, indicated polygenic control of this character.It was found that two non-genetic factors, day length and gibberellic acid (GA), may mimic the genetic factors for sex expression. Furthermore, evidence was presented, indicating that this sex controlling ability of GA and the modifying genes may be based on physiological conditions common to those two genetic and nongenetic factors.The overall mechanism of sex control in the cucumber and in other plants was discussed and a hypothetic model for the evolution of dioecism in flowering plants was proposed.     

Metabolic changes during rooting in stem cuttings of Casuarina equisetifolia L.: effects of auxin, the sex and the type of cutting on rooting

Rooting in terminal shoot and lateral shoot cuttings from 10-year-old elite trees of Casuarina equisetifolia L. in different sex groups was achieved after 20 days when the basal ends of the cuttings were dipped for 3 h in 20 ppm indole-3-butyric acid (IBA). Shoots derived from male plants rooted better than their female and monoecious counterparts, and the lateral shoots were more responsive to rooting than the terminal shoots. During rooting, the metabolic activities varied in both lateral shoot and terminal shoot cuttings derived from plants under different sex groups. Peroxidase and polyphenoloxidase activities were high during root initiation and showed a sharp decline thereafter. The polyphenoloxidase activity was higher in the lateral shoot than the terminal shoot cuttings. The rooted plantlets survived and established well in the field.Abbreviations IAA indole-3-acetic acid - IBA indole-3-butyric acid - NAA 1-naphthaleneacetic acid - PVP polyvinylpyrrolidone     

Identification of two markers linked to the sex locus in dioecious <Emphasis Type="Italic">Asparagus officinalis</Emphasis> plants

One hundred decamer primers of random-amplified polymorphic DNA were tested on dioecious Asparagus officinalis plants to identify sex-linked molecular markers. One primer (S368) produced two markers (S368-928 and S368-1178) in female plants. These two DNA markers were identified in 30 male and female plants, respectively, and a S368-928 marker was proved to be linked to the female sex locus. The female-linked S368-928 marker was sequenced and specific primers were synthesized to generate a 928 bp marker of sequence characterized amplified regions (SCAR) in female plants, SCAR₉₂₈. SCAR₉₂₈ could be used to correctly screen homozygous mm female plants of A. officinalis. However, results of Southern blot analysis suggest that the hybridization pattern of S368-928 was presented in both sex plants. This text was submitted by the authors in English.     

An in vitro method for screening for the presence of thepat-2 gene in tomatoes (Lycopersicon esculentum Mill.)

Under field conditions,pat-2, the gene which conditions parthenocarpy in tomatoes, is recessive. A simple method has been devised for distinguishing the heterozygote from the two homozygotes using tissue culture. Ovaries of plants segregating for thepat-2 gene were excised and cultured on a medium containing 100 ppm gibberellic acid. After three weeks in culture, three distinct ovary sizes could be seen. It was shown, using F ₃ progeny tests, that the largest ovaries corresponded to those plants homozygous for thepat-2 gene, the smallest ovaries corresponded to those plants homozygous for the wild type allele, and the intermediate sized ovaries were the heterozygotes. The ability to identify the heterozygote would greatly simplify a backcross breeding program aimed at incorporating thepat-2 gene into commercial cultivars by eliminating the need for an F ₃ progeny test to determine the genotype of a plant.Abbreviations GA ₃ gibberellic acid - IAA indole acetic acid - ppm parts per million     

Tests of a mechanistic model of one hormone regulating both sexes in Cucumis sativus (Cucurbitaceae)

A mechanistic model of sex determination in flowering plants, which assumes that one hormone has male and female cell receptors to inhibit one sex and induce the other independently, was tested in cucumbers (Cucumis sativus) by applications of hormones and their inhibitors. Applications of gibberellic acid (GA) and an inhibitor of its synthesis (paclobutrazol) showed that GA had dual effects on sex expression of promoting maleness and inhibiting femaleness. Conversely, applications of Ethrel (an ethylene release agent) and AgNO₃ (an ethylene action inhibitor) indicated that ethylene induced femaleness and inhibited maleness. Results of various combined applications of the two hormones and their inhibitors suggested that ethylene had overriding effects on GA and acted more directly on sex expression in cucumber. These experiments indicated that there is only one hormone, not two, regulating sex expression in cucumber, and that the sex hormone is likely to be ethylene. Results thus supported the assumption of the model that one hormone can regulate both sexes by inducing one and inhibiting the other independently. Model predictions were confirmed that from a monoecious line, female plants could be induced by increasing the ethylene level, and male plants induced by decreasing the ethylene level. On the other hand, as the model predicts, applications of ethylene or its inhibitor induced gynomonoecious, female, and trimonoecious plants from a hermaphroditic line. In conclusion, all assumptions and predictions of the one-hormone model were confirmed in the experiments.     

Modulation of root growth and organogenesis in thidiazuron-treated ginseng (Panax quinquefolium L.)

The morphoregulatory effect of thidiazuron (TDZ) applications to ginseng (Panax quinquefolium L.) seedlings and 3-year-old plants was determined. Applications of TDZ (0.22 and 2.20 ppm), either as foliar sprays or soil drench to greenhouse-grown seedlings over 18 weeks (2 weeks after sowing to 20 weeks when plants were harvested) induced similar responses, in particular, increased stem length and diameter, and shoot and root weight (economic yield). Single foliar applications of TDZ at 62.5 and 125 ppm to 3-year-old field-grown ginseng plants 3 months prior to harvest increased root biomass (economic yield) by 19 to 23%. Roots of TDZ-treated seedlings and 3-year-old field-grown plants developed thickened secondary roots on the upper part of the taproot. The root-like structure of these secondary roots was confirmed by histology. In addition, TDZ treatments induced adventitious buds on the shoulder of 3-year-old roots. These buds developed into shoots to give multi-stem plants following a period of dormancy, which was overcome with GA₃ (gibberellic acid) treatment prior to planting.Abbreviations TDZ = thidiazuron - GA₃ = gibberellic acid - BA = benzyladenine     

Effect of nurse cultures on the production of macro-calli and fertile plants from maize embryogenic suspension culture protoplasts

Fertile plants have been obtained from maize (Zea mays L.) embryogenic suspension culture protoplasts. Friable, embryogenic callus initiated from an immature embryo from a cross involving the genotypes A188, B73, and Black Mexican sweetcorn was used to establish a rapidly growing embryogenic suspension culture. After nine months in culture, high yields of viable protoplasts (30×10⁶/ gram fresh weight) were obtained following a 1.5 hour enzymatic digestion. Protoplasts cultured with feeder cells divided and formed embryogenic callus, from which male and female fertile plants were regenerated. Protoplast-derived R₁ plants were self-pollinated and immature R₂ embryos isolated for callus initiation. Female fertile plants have also been produced from protoplasts isolated from an R₂-derived suspension culture. Significant interactions between protoplast and feeder-cell lines were observed.Abbreviations BC backcross - BMS Black Mexican Sweetcorn - 2,4-D 2,4-dichlorophenoxyacetic acid - PWS protoplast wash solution (0.2 M mannitol, 80 mM CaCl₂) - FDA fluorescein diacetate - ABA abscisic acid     

谭清苏铁性别连锁的RAPD和SCAR分子标记

利用RAPD(Random amplified polymorphic DNA)分子标记技术,寻找谭清苏铁（Cycas tanqingii）中与性别相关的分子标记,筛选了160个10bp的随机引物,产生了2500多个RAPD条带。只有引物S0465 (CCCCGGTAAC)产生了一条大约500bp的雌性特异RAPD标记,该分子标记出现在所有的供试雌性植株中,而所有的供试雄性植株都不具有该标记。对该特异片段进行了克隆和序列测定,并根据序列分析结果将RAPD标记转化为重复性和特异性更好的特异特征序列扩增区域（SCAR）分子标记,并命名为STQC-S465-483。分子标记的建立可用于谭清苏铁幼苗性别的早期鉴定,为谭清苏铁就地保护和迁地保护提供技术支持。     

Effect of growth regulators and role of roots in sex expression in spinach

When 7-d-old plantlets of spinach (Spinacia oleracea L.) were immersed with their roots for 24 h in 25 mg/l gibberellic acid (GA₃), or 15 mg/l 6-benzylaminopurine (6-BAP), or 15 mg/l indole-3-acetic acid (IAA), or 10 mg/l abscisic acid (ABA) and subsequently grown on long (18-h) days, the ratio of plants with male and female flowers, which in the controls was almost 1:1 (48 and 52%, respectively), was greatly altered. The treatments with 6-BAP, IAA and ABA raised the percentage of female plants to 88, 76 and 71%, respectively; the GA₃ treatment increased the percent of male plants to 79%. When young, vegetative spinach plants (3 visible leaves) grown in 18-h days were cut a the root neck, and the shoots grown with their bases in nutrient solution, with adventitious roots either being allowed to develop or being systematically removed, 85% of the plants without roots became males, 85% of those with roots became females. But if the cut shoots were first, for 28 h, placed in a 15-mg/l 6-BAP solution and then grown in the absence of roots, the percent of female plants was restored to 84. These results fully agree with those obtained previously with hemp, namely, that plant growth regulators exert a regulating effect on the sex expression of dioecious plants when applied through the roots in early stages of development; that the root system plays an important role in determining the sex of these plants, that this role of the roots is associated with the synthesis of cytokinins in them. Dioecious short- and long-day plants do not differ in these respects.     

Pea aphid promotes amino acid metabolism both in Medicago truncatula and bacteriocytes to favor aphid population growth under elevated CO2

Rising atmospheric CO₂ levels can dilute the nitrogen (N) resource in plant tissue, which is disadvantageous to many herbivorous insects. Aphids appear to be an exception that warrants further study. The effects of elevated CO₂ (750 ppm vs. 390 ppm) were evaluated on N assimilation and transamination by two Medicago truncatula genotypes, a N‐fixing‐deficient mutant (dnf1) and its wild‐type control (Jemalong), with and without pea aphid (Acyrthosiphon pisum) infestation. Elevated CO₂ increased population abundance and feeding efficiency of aphids fed on Jemalong, but reduced those on dnf1. Without aphid infestation, elevated CO₂ increased photosynthetic rate, chlorophyll content, nodule number, biomass, and pod number for Jemalong, but only increased pod number and chlorophyll content for dnf1. Furthermore, aphid infested Jemalong plants had enhanced activities of N assimilation‐related enzymes (glutamine synthetase, Glutamate synthase) and transamination‐related enzymes (glutamate oxalate transaminase, glutamine phenylpyruvate transaminase), which presumably increased amino acid concentration in leaves and phloem sap under elevated CO₂. In contrast, aphid infested dnf1 plants had decreased activities of N assimilation‐related enzymes and transmination‐related enzymes and amino acid concentrations under elevated CO₂. Furthermore, elevated CO₂ up‐regulated expression of genes relevant to amino acid metabolism in bacteriocytes of aphids associated with Jemalong, but down‐regulated those associated with dnf1. Our results suggest that pea aphids actively elicit host responses that promote amino acid metabolism in both the host plant and in its bacteriocytes to favor the population growth of the aphid under elevated CO₂.     

Effect of gibberellic acid treatments, environmental conditions, and genetic background on the expression of theparthenocarpic fruit mutation in tomato

Summary Theparthenocarpic fruit (pat) allele causes a complex syndrome affecting different aspects of tomato reproductive development. This mutation affects stamen (reduced length and carpelloidy), ovule (arrested integument growth and unviability), and ovary (autonomous growth, i.e., parthenocarpy) development;pat mutant plants therefore have reduced male and female fertility. We studied the phenotypic expression patterns of thepat gene after treatments with gibberellic acid (GA₃) and under different growth seasons (late spring and autumn) and genetic backgrounds (backcross [BC] population after interspecific cross). GA₃ treatments were only effective in restoring carpelloid anthers to the wild-type phenotype. Compared to late spring, mutant plants grown in autumn had a lower frequency of carpelloid anthers and aberrant ovules and a higher seed set. Inflorescence position also affected thepat expression; upper inflorescences had low frequency of short anthers and aberrant ovules and an increased tendency to set seeds,pat expressivity was more variable in BC₁ plants segregating after interspecific cross withLycopersicon pennellii than in the originalL. esculentum line. Therefore, a role for minor genes that modify the quantitative expression of thepat mutation is postulated and discussed.Abbreviations ANOVA analysis of variance - BC backcross - GA gibberellic acid     

Environmental conditions affect sex expression in monoecious, but not in male and female plants of Urtica dioica

Urtica dioica is a sub-dioecious plant species, i.e. males and females coexist with monoecious individuals. Under standard conditions, seed sex ratio (SSR, fraction of males) was found to vary significantly among seed samples collected from female plants originating from the same population (0.05–0.76). As a first step, we investigated the extent to which SSR and sex expression of male, female, and monoecious individuals is influenced by external factors. We performed experiments to analyse: (1) whether the environment of a parental plant affects the sex ratio (SR) of its offspring, (2) whether SSR can be affected by environmental conditions before flowering, and (3) whether sex expression of male, female and monoecious plants that have already flowered can be modified by environmental conditions or by application of phyto-hormones. Within the range of our experimental design, SSR was not influenced by external factors, and gender in male and female plants was stable. However, sex expression in monoecious plants was found to be labile: flower sex ratio (FSR, fraction of male flowers) differed considerably between clones from the same individual within treatments, and increased toward 100% maleness under benign conditions. These results provide strong evidence that monoecious individuals are inconstant males, which alter FSR according to environmental circumstances. In contrast, we consider sex expression in male and female individuals to be solely genetically based. The observed variation in SSR between maternal parents cannot be explained by sex-by-environment interactions.     

Expression of P2X6, a purinergic receptor subunit,is affected by dietary zinc deficiency in rat hippocampus

The purpose of these experiments was to determine whether dietary zinc depletion affected protein expression in the hippocampus. Eleven weanling Sprague-Dawley male rats (21 d) were fed the AIN-93G diet containing 1.5 ppm zinc and supplemented with 30 ppm of zinc in the drinking water. After 1 wk, the rats were randomly divided into three groups: control (n=3), pair fed (n=3), and zinc restricted (n=5). All groups consumed the same diet. The zinc-restricted group consumed water containing no zinc. The rats were sacrificed 3 wk later. Chelatable zinc levels in the hippocampus, as measured by N-(6-methoxy-8-quinolyl)-para-toluenesulfonamide (TSQ) staining, were significantly reduced in the zinc-restricted group. Analysis of hippocampal protein expression by two-dimensional electrophoresis (2DE) revealed increased expression of the P2X₆ purinergic receptor in the zinc-restricted rats, as determined by MALDI mass spectrometry (MS) and database analysis. The data provided evidence for the dual effects of dietary zinc deficiency on the hippocampus, reducing ionic zinc levels and stimulating protein expression. The role the P2X₆ receptor plays in the physiological response of the hippocampus to zinc depletion remains to be determined.     

Effect of growth substances on flowering and sex expression in isolated apical buds of Spinacia oleracea

Apical buds of Spinacia oleracea L. cv. Matador were isolated from 7-day-old vegetative seedlings and grown in sterile culture under inductive long, or non-inductive short photoperiods. Flowering of isolated apices was inducible in long days in approximately 75% of the plants, and in short days by gibberellin treatment (about 40%) or by raising the temperature to 30–32°C (13%). In long days the percentage of flowering was further increased by gibberellin treatment (up to 90%), while it was unaffected by abscisic acid and was strongly decreased by Amo 1618 (55%). In long days the ratio of male to female plants was near 1. The percentage of female plants in long days was increased by gibberellins, and the percentage of male ones decreased by kinetin; as a consequence, the ratio of male to female plants was lowered to about 0.50 in both cases. Abscisic acid and Amo 1618 had the opposite effect, probably because they decreased the flowering in female plants, so that the sex ratio was shifted to 2.6 and 6.8, respectively. Simultaneous treatment with GA₃ reversed the effect of abscisic acid on the sex ratio, but the reversal of the shift to maleness induced by Amo 1618 was only partial. In short days, gibberellins also stimulated the flowering in female plants more than in male. However, when the flowering was induced by higher temperature, most flowering plants were male and kinetin increased their percentage further. The above results have been discussed in terms of different requirements for flowering in male and female plants.