Effect of Exogenous DNA on Acid Deoxyribonuclease Activity in Intact Roots of Vicia faba L.

Exogenous DNA has been administered to roots of Vicia faba inwhich this DNA stimulates an increase in acid deoxyribonucleaseactivity. This enzyme activity is associated with vesicles whichfollow whatappears to be the cytological pathway of foreignDNA in host tissues.     

Some Cytological Changes accompanying the Regeneration of Meristematic Activity at the Apex of Decapitated Roots of Vicia faba L.

The changes that took place in mitotic index (MI), labellingindex (LI) and the relative proportions of interphase nucleiwith different amounts of DNA have been investigated duringthe regeneration of meristematic activity at the apex of rootsof Vicia faba over the 144 h period following removal of thecap and apical mm of the meristem. Measurements were also madeof the corresponding changes that took place as cells were displacedbasally along the root from the apex over the experimental period.In both parts of the root, MI and the relative proportions ofnuclei with different DNA contents changed from levels similarto those at the apex of the controls at the start and end ofthe experiment to levels resembling those found in more matureparts of the root at 24 and 48 h. In contrast to these results,LI declined over the experimental period. These cytologicalchanges were aresult of the development of lateral root primordiain both the apical 2 mm of the decapitated roots and as cellswere displaced out of the meristem into more basal parts ofthe root. It was concluded that the events leading to the regenerationof meristematic activity at the apex of roots from which thecap and apical mm of the meristem were removed, are no differentfrom those which result in lateral formation as cells are displacedbasally along the primary root from the apex, and they takeplace over the same time interval in both systems.     

Cap Formation during the Elongation of Lateral Roots of Vicia faba L.

Cell proliferation was examined in the cap initials of newly-emerged0·2 and 4·0 cm long lateral roots of Vicia fabafrom an investigation of the passage of labelled cells throughmitosis following a 1-h pulse with tritiated thymidine. Celldoubling time was found to increase in this group of initialcells as the secondary roots elongated, this increase beinga result of a gradual lengthening in the duration of the mitoticcycle of both the fast and slow cycling meristematic cells andof a decrease in the size of both the growth fraction and thoseproliferating cells with a short cycle time. The rate of cellproduction by the cap initials was maximal in the newly emergedsecondary roots and showed a gradual decline with subsequentlateral elongation. This change in the rate of cell proliferationin the cap initials has been shown to be related to the initiationof the quiescent centre following lateral root emergence fromthe tissues of the primary. The number of cells making up the cap of 0·2–4·0cm long secondary roots is known to be constant and thus therate at which cells are sloughed off of the cap into the soilmust be the same as the rate of cap cell formation in theseroots, all of the cap cells being replaced every 6–9 days.The rate of cap cell formation in 0·2–4·0cm long secondary roots was used to calculate that one 11-dayold Vicia plant will have contributed between 56 000 and 85000 cap cells to the rhizosphere from its root system sincegermination.     

Cell Expansion during the Elongation of Lateral Roots of Vicia faba L.

Cell width, breadth, length, cross-sectional area, volume andthe ratio of the volume of the nucleus to that of the cell havebeen determined in the epidermis, cortex and stele over theapical mm of lateral roots of Vicia faba as they elongated fromjust-emerged to 4 cm in length. Cell volume increased basallyalong the root in each tissue, but this was not a result ofcell expansion taking place in all dimensions in the epidermis,cortex and stele. Thus, while increase in cell length, was themajor factor involved in cell volume increase basally alongthe root in the stele, the corresponding dimensions in the epidermisand cortex were cell width and cell breadth respectively. Cellvolume was greater in the cortex than in the other tissues andusually greater in the epidermis than in the stele. Cell lengthwas greater in the stele than in the other tissues, while cellbreadth was maximal in the cortex and cell width in the epidermis.Changes took place in the various measurements made as the rootselongated. These are discussed with respect to the onset oflateral root growth and to changes in the rate of growth ofthese roots as they elongate.     

Tritiated-thymidine Labelled Nuclei in Primordia and Newly-emerged Lateral Roots of Vicia faba L.

The utilization of ³H-TdR in DNA synthesis and the distributionof labelled nuclei have been determined in large primordia (LP)and newly emerged lateral roots (JE) of Vicia faba followingdifferent exposure periods. This nucleoside appears to reachLP and the more basal, unemerged parts of JE by way of the vasculartissue of the primary root, while those parts of JE which projectout from the epidermis of the primary obtain ³H-TdR directlyfrom the surrounding medium. The utilization of ³H-TdR in thesynthesis of DNA in basal segments of JE is also complicatedby the temporary quiescence of nuclei in this part of thesemeristems as LP elongate to form lateral roots.     

The Development of a Quiescent Centre in Lateral Roots of Vicia faba L.

During lateral root elongation there was a reduction in thenumber of cells present in the apical mm of the root, in thewidth of the root and in the number of rows of cells acrossthe root. At the same time, a quiescent centre, which was absentfrom large primordia and from lateral roots which had just emergedfrom the tissues of the primary root, was established. The quiescentcentre increased in size and cell number as the lateral rootelongated from 0·2 to 4 cm, the most rapid increase takingplace in roots between 0·2 and 0·5 cm in length.There was no correlation between root width and size of thequiescent centre in these roots. The cells in the quiescentcentre were smaller in size and had a lower labelling indexthan cells in other parts of the meristem. The distributionof labelled nuclei within the quiescent centrewas determinedand is discussed with respect to the site of root initial cells.     

Somatic embryogenesis in Vicia faba L.

The paper describes a method of somatic embryo induction in callus and suspension cultures of Vicia faba L. Callus was induced from immature cotyledons (green maturity stage) of white-flowering horse bean lines cultured on L2 medium (Phillips and Collins 1979) supplemented with 1% sucrose, 0.7% agar and different concentrations of 2,4-dichlorophenoxyacetic acid. The medium with 2.5 M 2,4-Dichlorophenoxyacetic acid was found optimum for embryogenic callus induction. Somatic embryos developed after transfer of the callus to media lower or zero 2,4-Dichlorophenoxyacetic acid and increased level of sucrose (2.5%). The release of somatic embryos from the callus was more apparent after transfer to liquid medium. There were various stages of somatic embryo development, i.e. globular, heart-shaped and torpedo ones.     

High Respiratory Activity of Guard Cell Protoplasts from Vicia faba L.

The rate of O₂ uptake was about 29 times higher in guard cellprotoplasts (GCPs) than in mesophyll protoplasts (MGPs) on aChi basis. The O₂ uptake was inhibited by respiratory inhibitors,but stimulated by respiratory uncouplers. On a Chi basis, theactivities of Cyt c oxidase and NADH-Cyt c reductase, mitochondrialenzymes, were about 27 and 35 times higher in GCPs than in MCPs.On a Chi basis, the ATP content was about 9 times higher inGCPs. The amount of ATP in GCPs was decreased by respiratoryinhibitors, an energy transfer inhibitor, and uncouplers ofoxidative phosphorylation. On a volume basis, GCPs had 8- to10-fold higher respiratory activities than MCPs, but had a lowChi content and lacked the activity of NADP-glyceraldehyde-3-phosphatedehydrogenase (NADP-GAPD), the Calvin cycle enzyme. From theseresults, we concluded that oxidative phosphorylation plays amain role in ATP production in guard cells and that guard cellshave a heterotrophic feature. Salicylhydroxamic acid (SHAM)in combination with KCN or NaN₃ strongly inhibited O₂ uptake,indicating the presence of cyanide-resistant respiration inguard cells. Phenylmercuric acetate (PMA), a potent inhibitorof stomatal opening, reduced the ATP content of GCPs by about90%, whereas it had a relatively small effect on the ATP levelof MCPs. The specific effect of PMA on GCPs is discussed. (Received March 24, 1983; Accepted June 8, 1983)     

Growth Substances in the Roots of Vicia faba

An investigation has been made into the growth regulators presentin ethanol extracts of the seedling roots of Vicia faba afterseparation on paper partition chromatograms, using segmentsof Avena coleoptiles and mesocotyls and of Pisum sativum.rootsas biological assay material. Acetonitrile purification shows the presence of at least threeauxins running in isobutanol: methanol: water, at Rfs of 0–0·25,0·4–0·6, and 0·65–0·95;the latter may represent two different auxins. A similar, butclearer, picture is shown by the ether-soluble acid fraction.Here an auxin at Rf 0–0·25 also stimulates rootgrowth and could be ‘accelerator ’. A second atRf 0–0·25 is an indole compound which inhibitsroot growth and does not seem to be be IAA. A third at Rf 0·8–1·0is also a root-growth inhibitor and gives no indole reaction.The ‘inhibitor ß’ complex was demonstrated(Rf 0·65–0·85) together with a number ofother inhibitors at lower Rf value. The ether-soluble neutral component also contains auxins orauxin precursors. The water-soluble, ether-insoluble fraction contains four readilyinterconvertible substances with auxin properties. They allappear to inhibit root growth and give no indole reaction.     

Wilting of Leaves in Vicia faba L.

Wilting of leaves of Vicia faba L., which occurs when the pressurepotential (_p) is zero, and the leaf-water potential () at wiltingboth depend entirely upon the solute potential at incipientplasmolysis (_so) and not on soil-water status. Wilting in V.faba is acropetal; this is consistent with the hypothesis thatthere is a gradient of decreasing _so up the plant and that wateris transferred from the lower to the upper leaves, hasteningthe overall water loss from the lower leaves to the point when_p is zero. The gradient in _so up the plant is of the order of3–8 bar. It is proposed that wilting when _p>0 (i.e. > _so) shouldbe ‘apparent wilting’ and that when _p0 (i.e. _so),‘true wilting’.     

A Cytological Analysis of the Antimetabolite Activity of 5-Hydroxyuracil in Vicia faba Roots

The effects of 5-hydroxyuracil (5-HU) (isobarbituric acid) upon cell elongation, mitosis, and DNA synthesis were studied in Vicia faba roots. 5-HU had no consistent effect upon root elongation. It blocked DNA synthesis (analyzed by photometric measurements of Feulgen dye in nuclei) during the first 6 hours of treatment; the block spontaneously disappeared by the 12th hour of treatment. Uracil and thymine had no effect upon this block of synthesis. Both thymidine and uridine reversed the block in 6 and 9 hours respectively. In all cases blockage of DNA synthesis was followed by inhibition of mitosis (determined by changes in the percentage of cells in mitosis) and resumption of DNA synthesis was followed by resumption of mitosis. Inhibition indices calculated from the mitotic data indicated a competitive relationship between 5-HU and thymidine and 5-HU and uridine. 5-HU is considered to block DNA synthesis by competing with thymidine for sites on enzymes involved in the synthesis. It is suggested that uridine reverses the block in synthesis by undergoing a conversion to thymidine.     

New Karyotypes of Vicia faba L.

Five new karyotypes of V. faba are presented and the types and positions of the structural changes combined in the construction of these new karyotypes are described. All their chromosomes are easily distinguishable by morphological criteria. These new chromosome complements are presently used to study the inter- and intrachromosomal distribution of induced chromatid aberrations and related problems.     

Cytoplasmic male sterility in Vicia faba L.

Summary In many higher plants, nucleo-cytoplasmic interactions lead to pollen abortion. In Vicia faba, cytoplasmic male sterility is unstable as the cytoplasm appears to shift from a sterile to a fertile state. In this report, five flower phenotypes are defined but the study is focussed on the progenies obtained from intermediate, semi-sterile plants with the same homozygous nuclear constitution during five successive generations. The results could be interpreted by quantitative modifications of at least four different kinds of cytoplasmic determinants.     

Intraspecific unilateral incompatibility in Vicia faba L.

Summary Unilateral incompatibility was discovered when crossing was attempted between different self-compatible types and subspecies of V. faba. Crossing in the direction female less self-fertile x male more self-fertile failed, whereas the reciprocal-crossing succeeded. Unilateral incompatibility developed with the evolution of less fertile and large seeded field bean types. How a cross fertilized (and self-incompatible) system may develop from a self fertilized one is discussed.The unilateral incompatibility in V. faba and other plant species is compared. The two-power competition hypothesis can explain all kinds of unilateral incompatibility reported so far in the literature. Breeding field beans for improved self-fertility is discussed.     

Lateral root formation in Vicia faba L.

MI was found to decrease in LP with increase in cell number, and reached minimal values just before the emergence of LP to form lateral roots. These changes in MI have been correlated with the accumulation of cells in G1, 24 hours before a lateral root is formed. The durations of C and the various phases of the mitotic cycle were also investigated in LP, and compared with those in small primordia. As lateral root primordia increase in cell number, the durations of C, S and G2 become longer, while G1 becomes shorter. Also MI and GF decrease while the proportion of quiescent cells increases. Thus, there is a gradual decrease in the rate of cell proliferation as primordia increase in size. The changes which take place in these parameters during lateral root primordium development have been compared with the events which occur in seed proliferating tissues at the onset of dormancy.     

Para-nitrophenyl Phosphate as a Substrate for Some Acid Phosphatases in Roots of Vicia faba

A cytochemical and biochemical study of acid phosphatases inroots of Vicia faba using p-nitrophenylphosphate(p-NPP) as substratehas shown the lack of specificity of the substrate which canbe acted upon by a K⁺-activated acyl phosphatase, nucleotidepyrophosphatase, glucox-6-phosphatase, 5' (3')-ribonucleotidephosphohydrolase, nucleotide phosph-transferase and those acidphosphatases demonstrable with ß-glycerophosphate.It is suggested that care should be taken in the interpretationof both biochemical and cytochemical studies employing sucha non-specific substrate as P-NPP. Vicia faba, broad bean, root, acid phosphatases     

Isozyme gene markers in Vicia faba L.

Summary This study was conducted to assess the genetic basis of the variability observed for the glutamate oxaloacetate transaminase (GOT), Superoxide dismutase (SOD), esterase (EST), and malate dehydrogenase (MDH) isozyme systems in different open-pollinated Vicia faba varieties. Individual plants showing contrasting zymogram patterns were simultaneously selfed and cross-combined. Crossing was unsuccessful in producing progeny, and only selfed progenies were suitable for genetical analysis of isozyme variability. Three zones of GOT activity were made visible. The isozyme of GOT-2 and GOT-3 zones were dimeric and under the control of three alleles at the Got-2 locus and two alleles at the Got-3 locus, respectively. The isozymes of the GOT-1 zone did not show any variability. Three zones of SOD isozyme activity were made visible. The isozymes occurring in the SOD-1 (chloroplastic isozyme form) and SOD-2 (cytosol isozyme form) zones were dimeric and under the control of two alleles at the Sod-1 and Sod-2 loci. The isozyme visualized in the SOD-3 zone (mitochondrial isozyme form) were tetrameric and under the control of two alleles at the Sod-3 locus. Apparently the isozymes made visible in the most anodal esterase zones EST-1, EST-2, and EST-3 were monomeric, and the occurrence of two alleles at each of two different loci explained the variability observed in the EST-2 and EST-3 zones. For MDH, only two five-banded zymogram pattern types were found, and every selfed progeny showed only one of the two zymogram type, indicating that each individual possessed fixed alleles at the loci controlling MDH isozyme. Got-2, Got-3, Sod-1, Sod-2, and Sod-3 appear to be five new isozyme gene markers that can be useful in Vicia faba breeding for linkage study, varietal fingerprinting, outcrossing rate estimate, and indirect selection for quantitative characters.     

An Analysis of Cell Proliferation in the Apical Meristem of Lateral Roots of Vicia faba L.

The relative proportions of the various proliferating and quiescentcells, cell doubling time, mean cycle time and the durationof the mitotic cycle and its various phases as measured fromthe passage of labelled cells through mitosis have been determinedfor the initial cells of the cap, epidermis, cortex and stele,for the epidermis together with the cortex and for the steleat various distances basal to the cap-quiescent centre boundaryin 1-cm long lateral roots of Vicia faba. Cell doubling timegenerally increased basally along these tissues as a resultof a gradual decrease in the size of the proliferating populationof cells. Cycle time of the fast-dividing cell population, however,was less between 750 and 800 µm basal to the cap-quiescentcentre boundary than in the corresponding initial cells largelyas a result of a decrease in the duration of G1, although changesalso took place in the durations of the other phases of themitotic cycle as cells were displaced basally along the root.From data reported in this paper as well as other results inthe literature, it appears that the proportions of quiescentcells arrested in G1 and G2 vary in the different groups ofinitial cells. Moreover, the proportion arrested in G1 appearsto decrease basally along each tissue, while that in G2 increases.     

Some Effects of Abscisic Acid and Water Stress on Stomata of Vicia faba L.

Vicia faba seedlings grown under a plastic tent in the laboratorywere either watered well throughout their growth period or weresubjected to a water stress treatment for several days priorto an experimental treatment. The effects of a further waterstress treatment or an application of an aqueous solution ofabscisic acid (ABA) on the stomata of these plants were determined.Stomata of previously water-stressed plants proved to be moresensitive than stomata of well watered plants to ABA appliedthrough the petiole via the transpiration stream and sprayedonto leaf surfaces. Stomata of previously water-stressed plantsclosed more rapidly and to a greater degree than stomata ofwell watered plants. The hormone had only a small effect whenapplied directly to epidermal fragments removed from both groupsof plants. Stomata of plants which had received a water stresspretreatment were less sensitive to a subsequent period of waterstress than were stomata of previously well watered plants.It is proposed that stomatal adaptation to water stress maybe related to changes in the hormonal balance of the plant.