Haploid Plantlet Formation from Female Gametophytes of Ephedra foliata Boiss. in vitro

Female gametophytes (at the archegonial stage) excised fromyoung ovules of Ephedra foliata Boiss, were cultured on a basalmedium (Murnshige and Skoog's combinations of major and minorsalts, Iron source, vitamins, myo-inositol along with 2 percent sucrose and 10 per cent coconut milk) under aseptic conditions.Growth and morphogenetic responses of the explants to auxinswere compared at different concentrations and a study of theirinteractions with cytokinins has also been made. At 2 mg 1^–1,2, 4-D induced profuse callusing which subsequently producedroots. NAA at 4 mg 1^–1 was optimal for callus growth androoting. Combinations of 2,4-D and kinetin were more effectivein inducing roots and shoot buds than those of 2,4-D and benzylamino-purine (BAP). Addition of BAP (0.05 mg ^1–1) to themedium containing optimal concentrations of NAA resulted information of a large number of roots. Kinetin induced only rootingin the presence of 4 mg 1^–1 NAA. A high concentrationof BAP (8 mg 1^–1), stimulated shoot bud formation. Forthe further development of shoot buds, neither auxin nor cytokininwas needed. Cytological observations revealed the presence ofhaploid number of chromosomes, i.e. seven. Ephedra foliata, tissue culture, callus, regeneration, 2,4-dichlorophenoxyacetic acid, naphthalene acetic acid, kinetin, benzyl amino-purine     

Epidermal structure and development of stomata in vegetative and floral organs ofHybanthus enneaspermus (Linn.)F. Muell

The present paper deals with the epidermal structure and ontogeny of stomata in vegetative and floral organs ofHybanthus enneaspermus. The epidermal cells are either polygonal or elongated with straight, sinuous or arched thick anticlinal walls. The surface of the cuticle shows parallel striations radiating from the guard cells or hair bases. Unicellular and uniseriate bicellular trichomes with verrucose margins have been observed on all organs. The mature stomata are anisocytic, paracytic, anomocytic and transitional between anisocytic and paracytic. The ontogeny of anisocytic and paracytic stomata is syndetocheilic or mesogenous, anomocytic is haplocheilic or perigenous, while that of the transitional type is mesoperigenous. Four types of stomata have been observed on all the vegetative and floral organs and their ontogeny from organ to organ of this plant is constant. Stoma with a single guard cell is the result of disintegration of one of the guard cells before or after pore formation. Contiguous stomata are also occasionally noticed.     

Ontogeny of Stomata on the Foliar and Floral Or{dot}gans of some Species of Crotalaria L.

The structure and development of stomata in eight species ofCrotalaria belonging to the family Papilionaceae are described.The study is mostly confined to the leaves but it has also beenmade on the floral organs of C. mysorensis, C. retusa, C. sericea,and C. triquetra. The stomata may be paracytic, anisocytic,anomocytic, diacytic, or with one subsidiary cell. The differenttypes occur individually or they are placed side by side evenon the same surface of an organ. In general the paracytic typeis by far the commonest, followed by anisocytic and anomocyticones. Diacytic stomata and those with one subsidiary cell arerelatively rare. Different types of stomata in various organsof the same plant develop mesogenously. The present investigationalso indicates that the inconstancy of stomata in the familyis due to (a) their diversity and (b) an increase in the numberof subsidiary cells either by their division or by the neighbouringperigenes becoming subsidiary cell-like.     

Structure and Development of Stomata on the Vegetative and Floral Organs of Some Amaryllidaceae

The structure and development of stomata in 10 Amaryllidaceousmembers are described. The study is made on the leaves and floralorgans of Amaryllis belladona, Crinum asiaticum, C. bulbispermum,C. latifolium, C. pratense, Pancratium sp., Polianthes tuberosa,and Zephyranthes atamsco, and on leaves of Agave sp. and Curculigoorchioides. In most of the organs of the different plants studiedmore than one type of stoma occur, even on the same surface.However, the stomata are largely anomocytic in Amaryllis, Crinum,Pancratium, Polianthes, and Zephyranthes, but they are tetracyticin Agave and Curculigo. An increase in the number of subsidiarycells in teracytic, tricytic, paracytic, and stomata with onesubsidiary cell has been noted. It may be due to wall formationin the subsidiary cells or by the neighbouring perigenes assumingtheir form. The range of variation in different types of stomatain each organ has been studied. Abnormal features such as juxtaposedor superimposed contiguous stomata, connections between guardcells of nearby stomata, transformation of meristemoids intoepidermal cells are observed. The development of different typesof stomata in different organs of the same plant is perigenous.The systematic position of different genera is discussed inlight of the present findings. A method of preparing epidermalimprints by the use of Depex, a mounting medium, and white ofan egg is also described.     

Stomatal responses of Vicia faba L. to indole acetic acid and abscisic acid

Evidence is presented that stomata in isolated epidermal peelsof Vicia faba L. open in darkness in response to the externalpresence of indole acetic acid (IAA) in the incubation medium.The effect of IAA is found to be overcome completely in thepresence of either TRIS or MES buffers. In the absence of buffer,V. faba stomata are shown to be influenced by IAA in a concentration-dependenttrend which reached a maximum at an [IAA] of 10^–3 molm^–3. Further investigations reveal that stomata in thisspecies can be shown to respond to the presence of IAA and anotherphytohormone, abscisic acid (ABA). IAA and ABA are demonstratedto be antagonistic in their effects provided the incubationconditions are suitable. The data are discussed in relationto stomatal responses of other species in different treatmentconditions. Recommendations are made with respect to standardizationof incubation media during epidermal peel experiments. Key words: Vicia faba, stomata, indole acetic acid, abscisic acid, buffers     

Development of Stomata in some Ophioglossum Species

This paper deals with the development of stomata in vegetativeand reproductive organs of five species of Ophioglossum. Thestomata are typically hap-locheilic or perigenous from organto organ. Superimposed twin stomata, stoma with aborted guardcells and persistent stomatal initial are seen. The presentstudies support the view of foliar nature of the spike of Ophioglossum.     

Structure and Development of Stomata on the Vegetative and Floral Organs in Some Members of Caesalpiniaceae

The structure and development of stomata in 19 species of thefamily Caesalpiniaceae are described. The study is mostly confinedto the leaves, but observations have also been made on othervegetative and floral organs of some species Stomata may beparacytic, anisocytic, anomocytic, and with one subsidiary cell.Occasionally a stoma is diacytic, cyclocytic, or actinocytic.Different types occur individually or may be found side by sideeven on the same surface of an organ. The most prevalent typein all the genera is paracytic except in Caesalpima where itis anomocytic. The development of an anomocytic stoma is perigenous,but those with subsidiary cells are largely mesogenous; rarelyparacytic stomata are mesoperigenous. In spite of diversityof stomata, different types of stomata have similar patternsof development in different organs of the same plant. The presentinvestigation also indicates that the inconstancy of stomatain the family is due to (a) their diversity and (b) an increasein the number of subsidiary cells either by their division orby the neighbouring perigenes becoming subsidiary cell-like.     

Arrangement of Stomata on the Leaves of Pelargonium zonale and Sedum stahlii

Stomata of Pelargonium zonale are orderly arranged on the surfaceof the lower epidermis. Inter-stomatal and Inter-trichome distancesas well as the number of sides of stomata, epidermal cells adjacentto stomata, and regular cells of the epidermis have characteristicprobability distributions. A model of epidermal developmentis constructed based upon the quantitative features of eachtype of cell. The primary restriction of the model is that stomainitials inhibit adjacent cells from becoming stoma initials. A second type of stomatal arrangement occurs in Sedum stahliiand an appropriate model for the ontogeny of epidermal cellsin this plant can also be constructed that has similar quantitativecharacteristics to those of real fields of cells. The applicationof modelling in plant morphology is discussed with respect tounderstanding ontogeny and phylogenic relationships.     

Structure and Development of Stomata in Vegetative and Floral Organs of Three Species of Kalanchoe

The structure and ontogeny of stomata in vegetative and floralorgans of three species of Kalanchoe is described. The matureanisocytic stomata are mono-cyclic or completely or incompletelyamphicyclic, rarely paracytic, transitional between paracyticand anisocytic and with a single subsidiary cell. The developmentof all the types is syndetocheilic or mesogenous from organto organ but the mature stomatal apparatus varies from organto organ as regards the number and arrangement of subsidiarycells. Abnormal stoma with a single guard cell and arresteddevelopment were observed on all organs. An abnormal stoma witha single guard cell develops directly from the meristemoid.     

Stomata and Structure of Tetraploid Apple Leaves cultured in Vitro

Leaves of anther-derived tetraploid apple (Malus pumila Mill.)shoots were examined by low-temperature scanning electron microscopy(LT-SEM). Leaves were serrate and wide with an undulating adaxialsurface due to convex epidermal cells, apparently without crystallineepicuticular wax. Stomata were absent from the adaxial surface,except for the marginal teeth which exhibited 40-60 stomataper leaf; they probably originated from residual mitotic activity.One third of abaxial stomata was occluded by the residual cuticleof the mother guard cell across the stomatal pore which rupturedwhen the stomata became functional. The stomatal index was 7·2(± 1·6) with 60-75 stomata mm^-2, i.e. abaxialstomata of tetraploid leaves expanded in vitro were less frequentthan those in triploid leaves either cultured in vitro (475-575stomata mm^-2) or grown on the tree (320-390 stomata mm^-2) wherethe stomatal index was 21 (± 4). Freeze-fracture transsectionsshowed that the tetraploid in vitro leaves were composed ofa layer of adaxial epidermal cells, followed by a single layerof palisade cells and four to five layers of spongy mesophyllcells and the abaxial layer of epidermal cells, in contrastto juvenile seedling-grown apple leaves in which the two layersof palisade cells comprised the majority (52-60%) of the leafvolume. The same morphological features, such as wide and lesspointed leaves, reduced stomatal density and stomatal index,and increased stomatal size that were previously reported fortree-grown tetraploid leaves were also expressed in vitro. Thus,causes of the stomatal deformation in tissue-cultured Rosaceaeare interpreted to be in part genetic and not purely environmental.Copyright1994, 1999 Academic Press Malus pumila Mill., apple, biotechnology, breeding, cryo-preservation, CO₂, juvenile, low temperature-scanning electron microscopy (LT-SEM), micropropagation, ploidy, stomata, tissue-culture, transpiration     

Some Observations on the Diversity of Stomata and Trichomes in Six Species of Dioscorea

Stomata and trichomes are described on the leaves of six speciesof Dioscorea. As many as six types were found in D. bulbiferaand D. oppositifolia, four types in D.hispida and D. wallichii,three types in D. belophylla, but only two types in D. alata.Although there is a diversity of stomata even on the same surface,the predominant type is anomocytic in all the species exceptD. bulbifera in which it is tricytic. Rarely a stoma is alsocyclocytic in D. bulbifera. An increase in the number of subsidiarycells in paracytic, tricytic, or diacytic stomata takes placeby the wall formation in them. Similarly a reduction in thenumber of subsidiary cells of a tetracytic stoma is the resultof lateral subsidiary cells assuming the form of epidermal cells.Abnormalities such as a stoma with one guard cell, degenerationof guard cells, and contiguous stomata are also met with. Theorganization of different types of stomata is studied in D.bulbifera and D. wallichii and shown to be perigenous. Capitateglandular hairs were seen on the leaves of D. belophylla, D.bulbifera, D. hispida, and D. wallichii but non-glandular, uniseriate,3-celled trichomes were observed only in D. hispida.     

Cellular Interactions in the Development of Stomatal Patterns in Vinca major L

The development of the leaf epidermis of Vinca major L. wasfollowed in situ by epi-illumination microscopy and evidencewas sought for cellular interactions. Stomata were often foundto be initiated in adjoining cells. The epidermal cells whichseparated such stomata when they had matured were formed fromthe same cells as the stomatal complexes themselves. The presenceof developing and mature stomata may influence only the orientationof divisions in neighbouring cells, and not the initiation andmaturation of stomata. There is great variability in the relativeorientation, timing and number of divisions which intervenebetween the first unequal division and the maturation of a stomaas well as the location of stomata relative to the spongy mesophylland minor veins. The results indicate that continuous short-rangeinteractions between the future guard cells and the adjoiningcells, rather than interactions between future stomata or afixed programme of development, are essential for the formationof the pattern of functional stomata in the mature leaf. Vinca major L., cell lineage, cellular interactions, development of stomata, epi-illumination microscopy, meristemoids, patterned differentiation, stomata     

Stomatal Pattern in Four Species of Monocotyledons

The distribution of stomata has been investigated in the leafepidermis of Chlorophytum comosum, Galanthus nivalis, Schizostyliscoccinea and Scilla lancifolia. The epidermis was consideredto consist of units of construction of two kinds: type A, along epidermal cell with a stoma at its distal end, and typeB, a long epidermal cell without an associated stoma. Exceptin Scilla, the probability of an epidermal unit being type Aincreases approximately with its length. Considering the epidermisas rows of units, alternating sequences of type A and type Bdo not occur randomly along the rows. In Chlorophytum, Galanthusand Schizostylis, both type A and B units tend to be aggregatedinto longer sequences than would be expected on a random basis.It is suggested that homoeogenetic induction (i.e. of like bylike) may be occurring during development. No case can be madefor homoeogenetic induction of units in Scilla. There is a slighttendency to periodicity of distribution of type A units in Galanthus,Schizostylis and Scilla, but this does not seem to representa primary element of pattern. There is interaction between rowsin the sense that unit ends (transverse walls) tend to avoidthose in neighbour rows; this affects the relative distributionof stomata, but there is no evidence of any direct interactionbetween stomata in different rows. Chlorophytum comosum, Galanthus nivalis, Schizostylis coccinea, Scilla lancifolia, leaf, epidermis, stomata, pattern     

Leaf‐morphology and leaf‐anatomy in Ephedra altissima Desf. (Ephedraceae,Gnetales) and their evolutionary relevance

Leaves in most extant Ephedra ‐species represent only rudimentary scales without chlorophyll. Photosynthesis is completely restricted to green shoots. Only some species, e.g. Ephedra altissima, develop folious leaves even when being mature. Morphology and anatomy of cotyledons, primary leaves, and subsequent folious leaves of Ephedra altissima were examined particularly with special focus on their vasculature. The results show that the reduction of the leaves is achieved by an extreme reduction of the adaxial leaf surface. Most parts of the leaf surface are therefore developed by the abaxial side. Apart from a reduced lamina, a thick cuticle and sunken stomata are the only xerothermic adaptations in leaves of Ephedra altissima. A hypodermis and endodermis are also not developed. The anatomical results indicate that ancestors of modern Ephedra‐ species might have evolved in a more humid climate, contrasting to the arid habitats in which extant Ephedra‐ species are native today. (© 2013 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Structure and Function in the Grass Ligule: Optical and Electron Microscopy of the Membranous Ligule of Lolium temulentum L.

Aspects of the structure and ultrastructure of the membranousligule of mature leaves of Lolium temulentum L. are described.In transverse section the ligule was lens-shaped and wedge-shapedin longitudinal section, 6 or 7 cells wide near the base and1 or 2 cells wide at the edges. Two uniseriate epidermes encloseda chlorenchymatous mesophyll tissue of varying thicknesses;both epidermes were continuous with the leaf adaxial epidermis.The cells comprising these three issues all appeared like typicalgrass epidermal long cells; elongate papillate cells were presentat the edges. No stomata, trichomes, intercellular spaces orvascular tissue were found in the ligule. A marked polarizationof ultrastructural complexity existed from the large-vacuolateabaxial epidermis to the ‘densely cytoplasmic’ small-vacuolateadaxial epidermis. Cells of the latter tissue contained numerousmitochondria, hypersecretory dictyosomes and abundant strandsof rough endoplasmic reticulum. Fluorescence microscopy providedevidence for the accumulation of a polysaccharide-containingmaterial within the periplasmic space next to the outer tangentialwall of adaxial epidermal cells. The ligule is considered tobe a highly organized and differentiated leaf organ with a pholosyntheticmesophyll and an adaxial epidermis active in the synthesis ofprotein and polysaccharide. Darnel, fluorescence microscopy, ligule, Lolium temulentum L., Poaceae, ultrastructure     

Organogenesis in the Cultured Female Gametophyte of Ephedra foliata

The female gametophyte of Ephedra foliata was used as an explantfor the production of haploids as it is composed of haploidcells, all of the same genotype. The regeneration of roots wasdependent upon the presence of NAA, while BAP had a modifyingeffect. At lower concentrations (0.05 parts 10^–6 and 3.5parts 10^–6) BAP enhanced the root promotion of NAA (0.05–4.0parts 10^–6). At higher concentrations of BAP (1–6parts 10^–6), roots and shoot buds were formed. Kinetinat 4.0 parts 10–6 with 0.5 parts 10–6 2, 4-D wasoptimal for shoot bud production in explants at the archegonialstage and 2, 4-D at 2.0 parts 10–6 with 0.5 parts 10–6kinetin was optimal for root formation. Cells of the callusand root tip had the haploid number of chromosomes, n = 7. Meristemoidswere located on the surface or embedded in the callus tissue.The deep seated meristemoids organized only root primordia,but the peripheral ones gave rise to root as well as shoot budprimordia. Initially, there was no vascular connection betweenthe shoot-bud and the callus. This was established later. Key words: Ephedra, Female gametophyte, Haploid, Tissue culture     

The Cuticle, Epidermis and Stomatal Ontogeny of Casuarina equisetifolia Forst

The cuticle, epidermis and stomatal ontogeny of Casuarina equisetifoliaForst. is described. The cuticle shows well marked impressionsof the epidermal cells and stomata. The epidermis of leaf andstem shows transversely oriented, tetracytic, mesoperigenousstomata with two lateral mesogene subsidiaries and two polarperigene neighbouring cells. Although the epidermal structureof Casuarina shows a good deal of resemblance with that of theBennettitales, it may not indicate any phylogenetic relationshipssince there are important differences in the structure and reproductionof the plants of these two groups.     

Tentoxin Suppresses Stomatal Opening by Inhibiting Photophosphorylation

Tentoxin and, to a lesser extent, dihydrotentoxin (both at 10mmol m^–3) reduce stomatal opening in epidermal stripsof Commelina communis in the light but not in darkness. Thiseffect was significantly greater in normal air than in CO₂-freeair. Fusicoccin overcame the tentoxin effect. However, tentoxindid not inhibit stomatal opening in the light in epidermal stripsof Paphiopedilum harrisianum, a species which lacks guard cellchloroplasts. It is concluded that tentoxin exerts its actionon stomata not by an ionophorous effect in the plasmalemma ofguard cells but by the inhibition of photophosphorylation intheir chloroplasts. The effects of DCMU and tentoxin on guardcells are discussed in terms of their effects on chloroplastsand the extent to which energy is supplied from this organelleduring stomatal opening in the light. The results indicate thatneither photophosphorylation nor non-cyclic electron transportin guard cell chloroplasts are essential for stomatal opening. Key words: Commelina, epidermal strips, Paphiopedilum, photophosphorylation, stomata, tentoxin     

Histological Changes of the Peanut (Arachis hypogaea) Gynophore and Fruit Surface During Development, and their Potential Significance for Nutrient Uptake

Histological changes in gynophores and fruits of Arachis hypogaeaL.cv. White Spanish were examined, utilizing scanning electronmicroscopy as well as light microscopy. The epidermis of theabove ground parts of gynophores is characterized by the presenceof stomata, lenticels and multicellular trichomes. Below groundportions of the same plant organ exhibit unicellular root-hair-likestructures. These protuberances of the epidermal cells can reacha very high density and length (up to 0.75mm) . Identical structurescan be found on the developing pod and are most prominent atthe reproductive stages R5-R6. In later developmental stagesthe hairs degenerate and the presence of large lenticels becomesthe obvious external feature of the pod. It is suggested thatthe substantial increase in surface area due to the hairs maywell be an anatomical adaptation for nutrient and water uptake. Arachis hypogaea, peanut fruit development, nutrient uptake     

Functional Stomata in a Variegated Leaf Chimera of Pelargonium zonale L. without Guard Cell Chloroplasts

Fluorescence microscopy indicated that chlorophyll was absentfrom epidermal and guard cells overlying all white areas andgreen areas (of certain leaves) in variegated leaves of Pelargoniumzonale, cv. Chelsea Gem. Stomata with chlorophyll-free guardcells, in general, responded normally to light and CO₂ as gaugedby direct measurements of stomatal aperture and by transpirationalwater loss studies, although stomata from white regions of variegatedleaves were more reluctant to open than stomata from green regionsof the leaves. Thus, functional stomata without guard cell chloroplastshave been discovered in another genus, namely Pelargonium, besidesthat originally discovered in Paphiopedilum. When stomata withchlorophyll-free guard cells opened, K⁺ accumulated in the guardcells. This indicates that chloroplasts are not essential forthe normal functioning of stomata and that the energy sourcefor driving stomatal movements can come from sources other thanphotophosphorylation. Key words: Guard cell chloroplasts, Leaf chimera, Pelargonium, Stomata