Mordanting Plant Tissues

Selected current ideas on the mordanting of plant tissues are presented in the hope that they will be of practical assistance, especially to the beginner. The nature of the mordanting process and the results which may be expected following mordanting are briefly described. Specific technics are presented for the mordanting of natural dyes, synthetic dyes generally, the basic synthetic dyes and the acid synthetic dyes.     

RNA extraction from plant tissues

Several protocols and commercial kits are used for the extraction of nucleic acids from different plant tissues. Although there are several procedures available to remove sugars, which hinder the extraction of clean genomic DNA, there are few to assist with extraction of RNA. Those presently used include precipitations with ethylene glycol monobutyl ether or lithium chloride (LiCl), or centrifugation in cesium chloride (CsCl) gradients, but these generally either do not allow high recovery of RNA, are time consuming, rely on hazardous chemicals or need special equipment. Here we present the use of the simple cation, Ca²⁺, which has been tested and shown to be very efficient for the precipitation of high molecular weight pectic sugars during RNA extraction. Results are presented for different plant tissues, especially tissues of peach and apple fruits at varying ripening stages.     

Heterogeneous growth of plant tissues

KORN, R., 1993. Heterogeneous growth of plant tissues. Heterogeneous growth is defined as different rates or patterns of growth in adjacent tissue regions, in contrast to homogeneous growth where a region expresses a uniform rate or pattern of growth. Heterogeneous growth is inspected in a variety of plant tissues and the pattern of expansion is characterized for each. In the case of epidermal cell proliferation, different growth rates for cell plates and old walls lead to the feature of coordinated growth in which slow growth of the former is compensated for by a faster rate of the latter. Examples include leaf epidermal cells above veins growing differently from those above areole regions, and pairs of guard cells of stomata ceasing to expand before other epidermal cells. In the alga Coleochaete only marginal walls grow, and at different rates around the colony, to generate a fractal, stochastic type of coordinated growth. In the fern gametophyte there are complex gradients of differential growth rates. Epidermal cells of apices are often of mixed growth, as cells at the summit undergo two dimensional expansion while cells along the flanks express one dimensional expansion. Coordinated growth requires matched rates where the constraining effect of the slower growing region is compensated for by a faster rate in an encircling region compared to the average rate of the overall tissue. Mixed and differential growth patterns do not necessarily create constraints and so lead to smooth tissue expansion. Emergence of some constraints leads to breaking of symmetry and disruptive growth as in the appearance of new axes found in organs and epidermal derivatives. In planar development heterogeneous growth appears to be the rule, and homogeneous growth the exception.     

A Mordanting Fixation for Intense Staining of Small Chromosomes

A combination iron-mordant fixative in which propionic acid is substituted for acetic acid has been found useful in preparing small plant chromosomes for carmine stained squashes. Propionic acid is better than acetic acid because it holds more iron in stable solution. The fixative is a 3:1 mixture of 95% alcohol and pure propionic acid which contains 400 mg. of Fe(OH)₃ per 100 ml. of propionic acid. The latter is previously prepared by dissolving the dry freshly prepared Fe(OH)₃ in it. To each 10 ml. vial of fixative is added a few drops of carmine stain. Standard aceto-carmine squashes of material fixed in this mixture show quick intense staining and are especially useful for differentiated chromosomes at mitotic prophase.     

Light-induced biophotonic emission from plant tissues

The emission of biophotons in the visible range, following a delay time of 2-200 seconds after exposure to light, has been measured in germinating seeds, roots, flowers, leaves, and cells. It was found that the biophotonic signals are reproducible and light-induced. The observed signals from germinating seeds of Phaseolus aures and decaying leaves of Eucalyptus are presented to show that the signals have characteristic kinetics and intensity. The kinetics of the signal was found to be independent of the stage of growth or decay, though its intensity varied with biological factors. The kinetics in the first minute is characterized by a single exponential decay term while that in the region t ≤ 200 s is characterized by two exponentials. The variation in the intensity of the signal with mass, state of hydration, and growth, and the effect of inhibitors in various systems (e.g. leaves, lichen, Chlorella) are reported.     

Calmodulin: localization in plant tissues

Calmodulin was purified from bovine brain by preparative SDS-polyacrylamide gel electrophoresis. The denatured, purified calmodulin was used to immunize rabbits to produce antiserum. This antiserum was used to study the distribution of calmodulin in plant tissues by indirect immunohistochemistry. The root tips from corn seeds, oat seeds, peanuts, spaghetti squash seeds, and the terminal buds of spinach were investigated. A method for plant tissue sectioning and inhibition of endogenous peroxide activity was developed. In the corn root section, reaction product from anti-calmodulin was found mainly in the root cap cells. Lesser but significant amounts of calmodulin were localized in metaxylem elements, in some stele cells surrounding metaxylem elements, in apical initials, and in the cortical cells. Similar findings were also observed in other root tips from oat seeds, peanuts, and spaghetti squash seeds. In the terminal buds of the spinach, calmodulin-stained cells were highly concentrated in the apical meristem and leaf primordium. These findings suggest that the high concentration of calmodulin in the root cap may be important in relation to gravitropism and growth development.     

Biogenesis of oxalate in plant tissues

Red beet root discs aerated in potassium phosphate for 2 to 3 days and young spinach leaves actively produce oxalate. A series of labeled compounds was supplied to each of these tissues to determine the extent of conversion to oxalate. Similar results were obtained with the 2 tissues except that in the leaf tissue glyoxylate and glycolate were outstandingly good precursors. Carbon from glucose, acetate, and particularly from some acids of the tricarboxylic acid cycle was recovered in oxalate. Extracts from both tissues were found to contain an enzyme which converts oxaloacetate to oxalate and acetate. The enzyme was partially purified and some of its properties are described. A pathway of oxalate synthesis which does not include glycolate or its oxidase is therefore proposed.