Root Tip Tissue Research Articles

Isozyme analysis of an active cassava germplasm bank collection

The genetic diversity among 28 accessions from the IPA (Pernambuco Agriculture Research Institute) Cassava Active Germplasm Bank was evaluated through the following isoenzymatic systems α,β-Esterase (EST), Peroxidase (POX), Glutamate Oxalacetic Transaminase (GOT) and Acid Phosphatase (ACP). Foliar and root tip tissues separated the accessions in 8 and 10 clusters, respectively. GOT and POX were the most polymorphic systems, resulting in 6 and 8 isoenzymatic patterns when foliar and root tip tissues were analyzed, respectively. The dendogram obtained by cluster analysis suggested that there was a similarity between leaf tissue and a morphological characterization indicating that highly inherited characters are good cassava descriptors. Based on the results obtained and those published earlier the enzymatic system could be standardized with the GOT and POX system being best suited for root tip and foliar tissue, respectively.

Morphological, agronomic, and isozyme characterization of some accessions of groundnuts (<i>Arachis hypogaea</i> L.)

Seventeen accessions of ground nuts (Arachis hypogaea L.) were characterized at the University of Ghana, Legon, 10 identity useful variation tor the genetic improvement of the crop. Using morphological and agronomic markers based on six qualitative and: 11 quantitative descriptors, wide variations were shown among the accessions. Each of the accessions was distinct except four, which fell into two groups of two each. Starch gel electrophoresis of crude protein extracted from the root-tip tissue of the accessions to study variation in esterase, peroxidase and acid phosphatase, also exposed variations among the accessions. Acid phosphatase showed the most variation while peroxidase showed the least variation. When all of the three isozymes were considered, it was possible to unambiguously distinguish 13 of the 17 accessions. The remaining four tell into two groups of two each. The results of the study indicated that isozymes are useful in the characterization of groundnuts.

Molecular characterization of plastidic phosphoserine aminotransferase in serine biosynthesis from Arabidopsis.

Serine biosynthesis in plants proceeds by two pathways; a photorespiratory pathway which is associated with photorespiration and a pathway from phosphoglycerate. A cDNA encoding plastidic phosphoserine aminotransferase (PSAT) which catalyzes the formation of phosphoserine from phosphohydroxypyruvate has been isolated from Arabidopsis thaliana. Genomic DNA blot analysis indicated that this enzyme is most probably encoded by a single gene and is mapped on the lower arm of chromosome 4. The deduced protein contains an N-terminal extension exhibiting the general features of a plastidic transit peptide, which was confirmed by subcellular organelle localization using GFP (green flourescence protein). Northern analysis indicated preferential expression of PSAT in roots of light-grown plants, supporting the idea that the phosphorylated pathway may play an important role in supplying the serine requirement of plants in non-green tissues. In situ hybridization analysis of PSAT revealed that the gene is generally expressed in all types of cells with a significantly higher amount in the meristem tissue of root tips.

Immunolocalization of β-(1→4) and β-(1→6)-D-galactan epitopes in the cell wall and Golgi stacks of developing flax root tissues

Most cell wall components are carbohydrate including the major matrix polysaccharides, pectins and hemicelluloses, and the arabinogalactan-protein proteoglycans. Both types of molecules are assembled in the Golgi apparatus and transported in secretory vesicles to the cell surface. We have employed antibodies specific to β-(1→6) and β-(1→4)-D-galactans, present in plant cell wall polysaccharides, in conjunction with immunofluorescence and electron microscopy to determine the location of the galactan-containing components in the cell wall and Golgi stacks of flax root tip tissues. Immunofluorescence data show that β-(1→4)-D-galactan epitopes are restricted to peripheral cells of the root cap. These epitopes are not expressed in meristematic and columella cells. In contrast, β-(1→6)-D-galactan epitopes are found in all cell types of flax roots. Immunogold labeling experiments show that both epitopes are specifically located within the wall immediately adjacent to the plasma membrane. They are also detected in Golgi cisternae and secretory vesicles, which indicates the involvement of the Golgi apparatus in their synthesis and transport. These findings demonstrate that the synthesis and localization of β-(1→4)-D-galactan epitopes are highly regulated in developing flax roots and that different β-linked D-galactans associated with cell wall polysaccharides are expressed in a cell type-specific manner.

Detection of β-tubulin in tomato seeds: Optimization of extraction and immunodetection

SDS-PAGE profiles of SDS-denaturated total protein (MODIL) extracts of imbibing tomato seeds showed no large qualitative differences from those of water-soluble (HEPES) proteins. However, the protein concentrations of the HEPES samples were significantly lower than the MODIL ones. The presence of SDS and DTT in MODIL buffer at an alkaline pH were essential for high yields of β-tubulin. The absence of DTT in HEPES extraction buffer may have allowed oxidation and subsequent loss of β-tubulin extracted from root tips of germinated seeds. Optimization of the β-tubulin extraction procedure and improvements of the immunodetection system enabled us to demonstrate the β-tubulin accumulation pattern in whole tomato seeds and in the different tissues. β-Tubulin accumulation in seeds was shown to be tissue-dependent, the highest concentration being found in embryo root tip tissue

Transformation of Chilli Pepper (Capsicum frutescens) with a Phenylalanine Ammonia-Lyase Gene

Hypocotyl explants of chilli pepper (Capsicum frutescens) were inoculated with Agrobacterium rhizogenes strain A13 harbouring the binary vector pBI121, which contains the CaMV 35S promoter linked to the GUS reporter gene. After 4 weeks of culture, the hairy roots produced were transferred onto Schenk and Hildebrandt (SH) medium supplemented with 250mgl-1 Cefotaxime. Subsequently they were selected in liquid SH medium containing 20mgl-1 kanamycin. When treated with X-glucuronide, vascular and root tip tissues were predominantly stained. A phenylalanine ammonia-lyase (PAL) cDNA from parsley (Petroselinum) was subsequently linked to the 35S promoter and transferred into hairy roots using the same A. rhizogenes strain. Successful transformation was confirmed by Southern hybridisation. Hairy roots containing the PAL transgene showed different PAL activity, slow growth and altered morphology.

Genetic Diversity among Plantagos XXXI. A Tertiary Trisomic of Plantago ovata. Nature, Behaviour and Origin.

A plant was isolated from the progeny of a cross between a translocation heterozygote mother and disomic pollen donor which carried an additional chromosome (2x+ 1) in all cells of the root tip and sporogenous tissue. The chromosome was unlike any in the standard chromosome complement. The nature and synaptic behaviour of this chromosome suggest that the aneuploid is a tertiary trisomic. The posible mode of origin of the extra chromosome and the aneuploid plant have been discussed.

Localization of Aluminum in Root Tip Tissues of Wheat Varieties Differing in Aluminum Tolerance

TitleLocalization of Aluminum in Root Tip Tissues of Wheat VarietiesDiffering in Aluminum ToleranceAuthor(s) Andrade, Leide R. M. de; Ikeda, Motoki; Ishizuka, JunjiCitation九州大学農学部紀要 ¦¦ 41(3/4) ¦¦ p151-156; Journal of the Faculty ofAgriculture, Kyushu University ¦¦ 41(3/4) ¦¦ p151-156Issue Date 1997-03URL http://hdl.handle.net/2324/24140Right

Spatial variation in the kinetics of aluminium (Al) uptake in roots of wheat ( Triticum aestivum L.) exhibiting differential resistance to Al. Evidence for metabolism-dependent exclusion of Al

Summary We have investigated the kinetics of Al uptake into various root segments and have compared uptake between two wheat (Triticum aestivum L.) cultivars that differ in their ability to resist Al toxicity. Following 60 and 180 min of exposure to Al, we found no differences in total Al accumulation between root tips (0–5 mm) and more mature root regions (5–10 and 10–20 mm). However, when roots were exposed to Al and subsequently desorbed in citric acid to remove exchangeable Al in the cell wall and in NH4Cl to remove the mucilage, non-exchangeable Al levels in root tips were 33 to 37% greater than in mature root regions. When the rate of accumulation of non-exchangeable Al in 20 mm root tips between an Al-resistant cultivar (PT 741) and an Al-sensitive cultivar (Neepawa) was compared, no differences were observed. When the same comparison was made in 5 mm root tips, the rate of Al accumulation in the Al-sensitive cultivar was nearly twice that in the Al-resistant cultivar. Furthermore, the rate of Al uptake in the Al-resistant cultivar increased dramatically in the presence of the metabolic inhibitor and protonophore, DNP, while DNP had little effect on uptake in the sensitive cultivar. This effect of DNP was also specific to root tip tissue. Our results support the hypothesis that DNP inhibits an Al exclusion mechanism operating at the plasma membrane in Al-resistant cultivars.

In vitro propagation of Phalaenopsis via culture of cytokinin-induced nodes

A new procedure for in vitro propagation of orchids belonging to the genus Phalaenopsis was developed. In contrast to commonly employed propagation methods that make use of leaf, root, or shoot tip tissues, we have used elongated stems of 6-benzyladenine-induced young seedlings as starting material for propagation. The elongated stem consisted of several nodes of which top nodes were used for cyclic propagation of new explants and the middle nodes for producing shoots or multiple adventitious buds. The whole procedure of proliferation could be completed within 7 months, and about 2,300 plantlets were produced from a single induced stem in a single year. This method may be used for propagation of seedlings in the case of lack of seeds in orchid breeding or for propagation of vegetative buds developed on flower stalks of rare orchid varieties when available flower stalks are limited. It may also have great potential for the propagation of wild threatened orchid species.

Changes in chromosomal ultrastructure during the cell cycle.

The surface structure of mitotic barley and rye chromosomes was studied by high-resolution scanning electron microscopy. Chromosomes with various degrees of chromatin condensation were prepared from untreated meristematic tissue of root tips. At lower magnifications the highly condensed chromosomes in metaphase and anaphase showed a compact structure with a smooth surface. The condensation starts from the centromeric region and the chromatids are often discernible in the still uncondensed telomeric region. Decondensation begins at the telomeric region during telophase. Parallel arrangement of fibres is a characteristic feature predominately seen in prophase and telophase chromosomes. Chromatin structures that resemble tiles on a roof or braided strands were often observed. Prophase and telophase chromosomes are particularly suitable for further studies of chromatin arrangement and organization in plant chromosomes.

Hardening of root cell walls: a growth inhibitory response to salinity stress

ABSTRACTPrimary roots of intact maize plants (Zea mays L.) grown for several days in nutrient solutions containing 100 mol m−3 NaCl and additional calcium, had relatively inhibited rates of elongation. Possible physical restraints underlying this salt induced inhibition were investigated. The inhibition did not involve reductions in osmotic potential gradients and turgor in the tip tissues responsible for root elongation growth. The apparent yield threshold pressure, which is related to capacity of cell walls to undergo loosening by stress relaxation, was estimated psychrometrically in excised root tips. Salinity increased yield threshold values. Comparative root extensibility values were obtained for intact plants by determining the initial (1 min) increase in root elongation rate induced by an 0.1 MPa osmotic jump. Comparative extensibility was significantly reduced in the salinized root tips. Salinity did not reduce capacities for water efflux and associated elastic contraction in root tip tissues of intact plants exposed to hypertonic mannitol. We conclude that cell wall hardening in the elongating root tips is an important component of root growth inhibition induced by long‐term salinization.

Intraplant Nuclear DNA Content Variation in Diploid Nuclei of Maize (Zea maysL.)

Diploid nuclei from stem, mesocotyl, nodal root and root tip tissue of two maize hybrids were examined with respect to their DNA content. The nuclei were isolated and stained with DAPI and passed through a flow cytometer-cell sorter. The titration curve for each tissue was determined. Significant variation was observed among nuclei of different tissue types. Stem and root tips had the highest diploid nuclear DNA amounts while 2-week-old mesocotyl had the lowest diploid nuclear DNA amount. These results provide evidence that during plant development and differentiation, the amount of DNA within a diploid nucleus changes through loss of specific DNA sequences. This study also demonstrates the sensitivity of flow cytometry in detecting small intraplant variation in nuclear DNA.

Morphological and isozyme characterisation of Arachis pintoi Krap. et Greg. nom. nud. germplasm

Arachis pintoi, a wild perennial relative of the peanut, originated in Brazil, is of increasing importance to pasture improvement in the tropics. Its available genetic base, however, is very narrow, and intensive collection is being carried out. This study aimed to characterise the existing eight accessions in the CIAT germplasm collection in order to describe the existing variation, identify plant types, and discover possible duplicates. Morphological characterisation, based on 60 morphological descriptors, demonstrated that variation exists. Numerical taxonomic techniques were used to rank the importance of these descriptors. Cluster analysis, conducted on those pre-selected morphological characters, grouped the accessions into two major plant types: one homogenous, and the other falling into four distinct subgroups. Polyacrylamide gel electrophoresis (PAGE) was applied to root-tip tissue. Isozyme characterisation by PAGE showed polymorphisms in the isozymes α-EST, ACP, GOT, and DIA. When analysed together, the four isozymes could uniquely distinguish every accession of A. pintoi held in the CIAT germplasm collection. Correspondence and cluster analyses of these isozyme patterns showed the morphologically homogenous plant type to be heterogenous, and the heterogenous morphotype to have similar banding patterns. The lack of continuous patterns of variation in both morphology and isozymes manifests the need for further germplasm collection of A. pintoi.

Chromosome morphology and number in various populations of Hydrilla verticillata (L.f.) Royle

Hydrilla verticillata (L.f.) Royle root tip karyotypes were not different among 20 populations from Australia, China, India, Malaysia, Panama and the USA, and the combined karyotype derived from these populations was similar to that previously reported. Root tip tissue was sometimes endopolyploid, with diploid (2 n = 2 x = 16), triploid (2 n = 3 x = 24) and tetraploid ( 2n = 4x = 32) cells observed in the same populations, and diploid and triploid cells often observed in the same root tip. Diploid cells only were observed in root tips from a population of Panang Island. Hydrilla is probably diploid because diploid cells were observed in all populations (except Beijing, China, which was predominantly tetraploid). Therefore, there is no apparent reason why all populations should not be sexually compatible, and atleast partially fertile, based on chromosome make-up.

Physical restraints underlying short-term inhibition by auxin of root elongation in intact maize seedlings

This report investigates physical changes associated with the short-term inhibition of root elongation in intact maize seedlings (Zea mays L. vs. Halamish) by exogenous auxin. Movement of root tips was assayed by video microscopy in control roots, roots grown for 45 min in 10−6 M indole3-acetic acid (IAA), or roots chilled for 3 min at 11°C. IAA and chilling treatments similarly reduced root elongation rates (from 29 ± 6 μm min−1 to 6 ± 2 μm min−1). Initial rates of root tip contraction induced by 300 mOsmol mannitol were used to calculate tissue contractibility values. These allowed a comparison of effects of IAA and chilling treatments on apparent rates of water transport out of the root tip tissues. Chilling treatment reduced root tip contractibility by 66%, whereas IAA had much less effect (26% reduction). Roots were also exposed to an osmotic jump treatment; the initial osmotically induced increase in elongation rate was used to determine root tip extensibility values. Both IAA and chilling treatments reduced root tip extensibilities by 57%. Inhibition of wall-yielding properties, rather than hydraulic limitations, appeared to be primarily associated with inhibition of intact root tip elongation by exogenous IAA.

Timing, magnitude, and impact of acidic deposition on sensitive forest sites

Adverse effects of acidic deposition on forest health are most likely to occur in forests which develop a thick raw or “mor” humus layer in which the effective cation exchange capacity is highly sensitive to acid input. A study of the trend of exchangeable Ca and Mg ions in sensitive humus layers over the past six decades indicated that a downward shift in equilibrium has occurred that is consistent with theories of ion mobilization and coincident in time with increasing acidic deposition in the mid-1900's. Independent records of a base cation mobilization in wood supports the view of a change in the root zone in sensitive forest sites and in lake water chemistry. Induced nutrient deficiency in declining forest stands was indicated by a high Al∶Ca ratio in fine root tip tissue as a marker of altered uptake efficiency which leads to increased vulnerability to biotic factors and greater sensitivity to abiotic stresses. Evidence indicates that sensitive sites were damaged by acidic deposition 20 to 40 yr ago, long before the problem of “acid rain” was recognized.

The Level of Abscisic Acid in the Root Tip of Runner Bean Seedlings: Implications for the ABA-Uptake Carrier

Summary High performance liquid chromatography and gas chromatography using an electron-capture detector have been combined in a method for the determination of the content of free abscisic acid in small amounts ( Phaseolus coccineus L. cv. ‹Prizewinner› root-tip tissue. The identification of abscisic acid was verified by mass spectrography and the method gave estimated overall recoveries of abscisic acid from the tissue consistently greater than 60%. A stereological analysis of transmission electron micrographs of thin sections was used to determine the relative volumes of cellular compartments at either end of that region of the root tip in which carrier-mediated abscisic acid transport has been shown (Astle, M. C. and P. H. RUBERY (1980) Planta, 150 ,312–320). Using the two sets of data and published estimates of the pH of cytoplasmic compartments, the concentration of abscisic acid in the major compartments was calculated. The results define (for unstressed seedlings as used by Astle and Rubery) the range of concentration of abscisic acid occurring in the cytoplasm and cell wall compartments in the region of the root in which the carrier was detected. The values for abscisic acid concentration in the cell wall compartment are so low relative to the K m of the transport carrier that it is not reasonable to suppose that the presence of the carrier will affect the distribution of abscisic acid between compartments to any appreciable extent under normal conditions.

The effect of aluminium on uptake and distribution of magnesium and calcium in roots of mycorrhizal Norway spruce seedlings

Spruce seedlings (Picea abies (L.) Karst.) colonized with Lactarius rufus (Scop.) Fr. or Lactarius theiogalus Fr. were grown in an axenic silica sand culture system with frequently renewed nutrient solution. After successful mycorrhizal colonization the seedlings were exposed to 800 μM Al(NO3)3 (pH 3.9) for 13 or 17 weeks. Concentrations of Al, Mg, and Ca in the tissues of the mycorrhizal root tips were determined by X‐ray microanalysis. After 13 or 17 weeks of exposure to Al, high Al concentrations were found in cell walls of all mycorrhizal tissues except the stele tissues. Compared to the controls Mg levels in most of the mycorrhizal structures were reduced by Al treatment. Calcium levels in cortex cell walls of root tips colonized with L. rufus were reduced by exposure to Al. However, in cell walls of the stele Ca levels were significantly increased. No differences in Al or Mg levels were detected in structures of mycorrhizal and non‐mycorrhizal root tips from the same individual seedlings. These results suggest that (1) the endodermis is the primary barrier to radial Al transport and (2) the presence of a hyphal sheath did not prevent Al from reaching the root cortex and from displacing Mg and Ca.

High-performance liquid chromatography of deoxyribonucleoside di- and triphosphates in tomato roots

The levels of endogenous deoxyribonucleotides in plants are greatly affected by the absence of the micronutrient boron. Described here is a high-performance liquid chromatographic assay for quantitating deoxyribonucleoside di- and triphosphates in tomato root tip tissues under boron-sufficient and boron-deficient conditions. The extraction procedure consists of (a) removing phenols from the tomato root tips by grinding frozen tissue in 6% trichloroacetic acid containing polyvinylpolypyrrolidone, (b) neutralizing the extract with freon-amine, (c) selective degradation of ribonucleotides with periodate, followed by (d) treatment with rhamnose and methylamine prior to injection of the deoxyribonucleotides into an anion-exchange column. Results indicate that the eight deoxyribonucleoside di- and triphosphates can be reproducibly resolved and quantified. By this extraction procedure, column life was extended to more than 70 runs per cartridge. This assay technique can be used with less than 500 mg of fresh root tip tissue and allows quantitation of plant deoxyribonucleotides in the picomole range.