Root Formation in Pea Cuttings

Auxin was applied to the upper part of the cuttings, which were both decapitated and disbudded on the same day. The applied auxin was removed by redecapitating the cuttings at different time intervals. In a second experiment, auxin was applied either to the upper or lower part of the decapitated and disbudded cuttings at different time intervals. In cuttings, which were redecapitated after 1 and 2 days, the root formation was reduced considerably. The redecapitation after 3 days had no adverse effect on the root formation. Cuttings treated with auxin at different time intervals showed a weaker root promotion on days 0 and 1 than on the subsequent days. The results emphasize the fact that auxin is active only during the first part of the root initiation phase. A continuous flow of auxin for a period of the first 3 days during the root initiation is of overriding importance. There appears to be at least two different stages of the root initiation phase, (ia) auxin active stage, and (ib) auxin inactive stage. The results also seem to indicate that some other factors, in addition to auxin, are active during the first stage of the root initiation phase.     

Root formation in pea cuttings: Effects of combined application of auxin and cytokinin at different developmental stages

Cuttings of pea cv. Alaska and ov. Kelwo were both decapitated and disbudded at different time intervals after cutting. Auxin and cytokinin combined in different ratios were applied to the upper part of the decapitated and disbudded cuttings. The effects of different ratios of auxin and cytokinin were not the same when applied at different developmental stages of the root initiation phase. The results seem to demonstrate an interaction between auxin and cytokinin at different ratios throughout the root initiation phase. The effects of combined application of auxin and cytokinin suggest that different stages of the root initiation phase require different levels of auxin and cytokinin. A higher level of auxin and either lower or equal level of cytokinin may be needed only in the early stages. During the subsequent stages a lower level of auxin in combination with a higher level of cytokinin seems to be more conducive.     

Root Formation in Pea Cuttings III.

Cuttings were either decapitated or both decapitated and disbudded at different time intervals. Cytokinin, at different concentrations, was applied to the cuttings in lanoline. Higher concentrations of cytokinin inhibited root initiation during the early stage. However, the inhibitory effect of cytokinin disappeared during the later stage of root initiation. Lower concentrations of cytokinin promoted the root initiation during the early stage. This effect was observed on cuttings which were only decapitated. These results seem to indicate that the influence of cytokinin changes with the stage of development. There seems to be an interaction between cytokinin and one or more other growth factors. A possible reason for this may be that cytokinin, in higher concentrations, produces inhibitory effects during the early part of root initiation by blocking the activity of auxin. The loss of the inhibitory effect of cytokinin during the later part of the initiation phase suggests that, at this stage, developing root primordia are capable of controlling the level of active cytokinin and thus do not react to the exogenous application of cytokinin.     

Polyamines and Adventitious Root Formation in the Juvenile and Mature Phase of English Ivy

The involvement of polyamines during adventitious root formationwas evaluated using a de-bladed petiole rooting assay for theeasy-to-root juvenile and difficult-to-root mature phase ofEnglish ivy (Hedera helix L.). Auxin (NAA 0.1 mM) stimulatedroot formation in juvenile phase cuttings, but failed to promoterooting in the mature phase. The addition of putrescine, spermineor spennidine (1.0 mM) with or without NAA (0.1 mM) did notaffect the rooting response in either the juvenile or maturephase cuttings. There was a significant increase in endogenouslevels of putrescine and spermidine in NAA-treated cuttings,but the only significant difference between the root formingjuvenile and the non-root forming mature phase cuttings wasan increase in putrescine levels. In NAA-treated juvenile cuttings,the polyamine biosynthesis inhibitor DFMA (1.0 mM) promotedroot formation from 9.2 to 14.5 roots per cutting, while DFMO(1.0 mM) reduced root formation from 9.1 to 1.4 roots per cutting.The promotion of rooting by DFMA was completely reversed byputrescine (1.0 mM), but putrescine, spermine or spermidine(1.0 mM) could not reverse the inhibitory effect of DFMO. NeitherDFMA nor DFMO promoted root formation in mature phase cuttings.DFMA was also added to NAA-treated juvenile petioles at variousstages during the root formation process. DFMA promoted rootingwhen applied during the early stages of root induction (0–3d), but became inhibitory to root formation when applied duringthe organization (6–9 d) or root elongation stages (9–12d). Key words: Hedera helix, organogenesis, root initiation, polyamines, DFMA, DFMO     

The nature of the dual effect of auxin on root formation in Azukia cuttings

In disbudded Azukia stem cuttings, auxin exerted a dual effecton root formation. The first phase of auxin action is identifiedwith the acceleration of cell division, especially longitudinaldivision. In cuttings treated with auxin during the first 24hr, longitudinally divided cells were observed in all 12 rootprimordia, while in water-treated cuttings, such cells wereobserved only in 8 root primordia. The second phase is the promotionof the reaction in which root primordia unable to develop furtherwithout auxin supply develop into roots. Irrespective of thetreatment during the first 24 hr, the auxin-treatment duringthe second 24 hr increased the number of roots protruding fromthe cuttings. Portulal applied during the first 24 hr increased the numberof root primordia which contained longitudinally divided cells.Gibberellin applied during the first 24 hr inhibited both transverseand longitudinal divisions in root primordia. ¹ Supported in part by Grant No. 139011 from the Ministry ofEducation, Japan. ² Present address: Junior College of Toyo University, Hakusan,Bunkyo-ku, Tokyo 112, Japan. (Received June 13, 1978; )     

Influence of ethylene on adventitious root formation in mung bean hypocotyl cuttings

The relationship between ethylene and adventitious root formation in mung bean hypocotyl cuttings was studied.Ethephon, an ethylene-releasing compound, at 5 x 10 -5 M increased root number and root dry weight on hypo-cotyl cuttings. When ethephon was applied to hypocotyl at different times after excision, there were two effectivetimes for root production i.e. between 06 h and 18-24 h. These two time periods correspond to the induction phase and the late initiation phase of root development, respectively. After excision, three peaks of ethylene productionwere observed. The first peak commencing at 6 h started the sequence of reactions leading root formation, the second peak appearing at 12 h coincided with the beginning of the increase of the IAA level during primordia initiation, and the third peak showing at 48 h played a role in root differentiation and growth. Ethylene stimulated rooting by enhancing the increase in auxins. Thus it appears that the IAA-induced ethylene production may be a factor involved in the stimulation of adventitious root formation.     

Auxins or Sugars: What Makes the Difference in the Adventitious Rooting of Stored Carnation Cuttings?

Cold storage of cuttings is frequently applied in the vegetative propagation of ornamental plants. Dianthus caryophyllus was used to study the limiting influences of auxin and sugars on adventitious root formation (ARF) in cuttings stored at 5°C. Carbohydrate levels during storage were modulated by exposing cuttings to low light or darkness. The resulting cuttings were treated (or not) with auxin and planted, and then ARF was evaluated. Carbohydrate levels in the cuttings were monitored and the influence of light treatment on indole-3-acetic acid (IAA) and zeatin (Z) in the basal stem was investigated. Dark storage for up to 4 weeks increased the percentage of early rooted cuttings and the final number and length of adventitious roots, despite decreased sugar levels in the stem base. Light during cold storage greatly enhanced sugar levels, particularly in the stem base where the Z/IAA ratio was higher and ARF was lower than observed in the corresponding dark-stored cuttings. Sugar levels in nonstored and dark-stored cuttings increased during the rooting period, and auxin application enhanced the accumulation of sugars in the stem base of nonstored cuttings. Auxin stimulated ARF most strongly in nonstored, less so in light-stored, and only marginally in dark-stored cuttings. A model of auxin-sugar interactions in ARF in carnation is proposed: cold storage brings forward root induction and sink establishment, both of which are promoted by the accumulation of auxin but not of sugars, whereas high levels of sugars and probably also of cytokinins act as inhibitors. Subsequent root differentiation and growth depend on current photosynthesis.     

Variation in the lag phase and uniformity of the length of the rooting phase prior to adventitious root production in cuttings of Agathis australis

Cuttings of Agathis australis (D. Don) Lindl passed through a well-defined series of morphological changes prior to root emergence. These phases were incorporated into a morphological index which can be used as a guide for the selection of cuttings at known developmental and anatomical stages. After a variable period (lag phase) during which no external change occurred there was an increase in stem diameter a few milimetres above the cut base. This swelling gradually increased in size and isolated bulges developed. Longitudinal splits then arose in the epidermis over the bulges, followed by root emergence through the splits. Root initiation occurred shortly after the sub-basal swelling commenced in cuttings that eventually rooted. Removal of the basal 8 mm of a rooted cutting (which included the roots) usually led to re-rooting of the cuttings. However, if the roots were merely trimmed off, the cutting never formed new roots and always died. The basal region apparently has the capacity to produce only one set of roots. Occasionally the stem diameter continued to increase and the swelling extended to include the basal region. Such cuttings never formed isolated wellings and never rooted.
In general the younger the plant from which the cutting was taken, the shorter the lag phase and the higher the final percentage rooting. Cuttings taken from older plants had a lower rooting percentage and a more variable lag phase, which was related to the time of year the cuttings were taken since root emergence always occurred in spring. Irrespective of the age of the original material there was a constant time period (3–4 weeks) from root initiation to root emergence.     

Seasonal Changes in Auxin Effect on Rooting of Douglas-Fir Stem Cuttings as Related to Bud Activity

The relation of seasonal bud activity to the periodicity of rooting in Douglas-fir, Pseudotsuga menziesii (Mirb.) Franco, stem cuttings was studied in combination with auxin and cold storage treatments. Cuttings were collected in all months except April and May, for 3 years. Rooting was least in September and October when bud dormancy was most pronounced, greatest in December and January if exogenous auxin was applied, or in February and March if no auxin was used. The buds contributed significantly to rooting from January to April, and were responsible for differences in rooting of terminal and lateral cuttings during this period. Auxin did not enhance rooting in September and October, but at other times it replaced or supplemented the role of vegetative buds in promoting rooting. Auxin also removed the differences in rooting between lateral and terminal cuttings. Cold treatment in October and November removed bud dormancy and enhanced rooting. After November the need for auxin or cold treatment diminished and rooting without either treatment reached a maximum in February and March. Auxin did not change the seasonal pattern of rooting but broadened and enhanced the rooting response in favor of earliness. These results are discussed in relation to the effect of bud activity on auxin response and root initiation. The hypotheses are proposed that cambial dormancy or auxin deficiency is not the limiting factor during bud dormancy, and that cold treatments have the effect of bringing inhibitors and promoters into proper balance for optimum rooting response.     

Effect of Main Root Decapitation on Root Branching in Flax Seedlings

Root branching patterns in intact and decapitated flax (Linum usitatissimumL.) roots were compared. The number of initiated primordia in the control and decapitated roots was similar, but decapitated roots produced an increased number of lateral roots owing to an increase in the number of primordia developed into the laterals. It is suggested that the apical meristem influences lateral root development only at the stage of root emergence from the parent root.     

Influence of Cycloheximide and Actinomycin D on Initiation and Early Development of Adventitious Roots

From leaf cuttings of the bean Phaseolus vulgaris L. adventitious roots form on the petiole. This root formation is stimulated by treatment with auxin. Simultaneous or subsequent application of cycloheximide irreversibly inhibited dedifferentiation, so that root production was completely prevented. The effects of actinomycin D application depended upon the stage of development of the root primordium. Cells in the first stage of dedifferentiation were extremely sensitive. When actinomycin D was applied later than 6 h after cutting, its inhibiting effect gradually diminished. It is concluded that an actinomycin D-sensitive process occurring early in dedifferentiation is crucial for root initiation. A second, less actinomycin D-sensitive process occurring later in dedifferentiation is required for the further development of the root primordium. During the initiation and development of the root primordium protein synthesis is required.     

Inhibition of Rooting of Cuttings by Gibberellic Acid: With one Figure in the Text

Gibberellic acid inhibited rooting but increased extension ofpea and bean stem cuttings. Indolylacetic acid usually had acontrary effect both on extension and on root initiation. Itwas found possible to separate the effects of gibberellic acidon extension and rooting. Small doses of gibberellic acid appliedto the bases of cuttings reduced rooting without increasingextension. Small apical doses increased extension without reducingrooting. Disbudding cuttings prevented extension, but gibberellicacid inhibited rooting of disbudded cuttings as strongly asthat of normal cuttings. It has thus been found necessary todiscard an earlier hypothesis that inhibition of rooting bygibberellic acid was a consequence of a diversion of essentialmetabolites to the extending apical regions of the cutting.The antagonism between gibberellic acid and indolylacetic acidwas non-competitive. Gibberellin A₁, known to occur naturallyin Phaseolus, was as active a rooting inhibitor as gibberellicacid. It is now believed that inhibition of rooting of cuttingsby gibberellins is a direct local effect, preventing those earlycell divisions involved in transformation of mature stem tissuesto a meristematic condition.     

Activity of some glycolytic and pentose phosphate pathway enzymes during the development of adventitious roots

Activities of phosphofructokinase (PFK, EC 2.7.1.11), glyceraldehyde 3-phosphate (NAD) dehydrogenase [G-3-PD(NAD), EC 1.2.1.12], glucose 6-phosphate dehydrogenase (G-6-PD, EC 1.1.1.49), and 6-phosphogluconate dehydrogenase (6-PGD, EC 1.1.1.44) were determined in bean cuttings (Phaseolus vulgaris L. cv. Top Crop) over 4 days, encompassing adventitious root primordium initiation and development. Effects of applied auxin and “endogenous root-forming stimulus”(ERS) on enzyme activities, concentrations of reducing sugars, and primordium development were also determined during the first 4 days of propagation. Effects of auxin were determined through use of applied indole-3-acetic acid (IAA) or 2,3,5-triiodobenzoic acid. Effects of ERS were evaluated by means of decapitation of cuttings. Increased basipetal transport and increased metabolism of reducing sugars occurred in leafy cuttings in response to applied IAA and to ERS. Primordium development and activities of the four enzymes increased in leafy cuttings under conditions that simultaneously increased basipetal transport and metabolism of reducing sugars. Three types of enzyme activity response were found: (i) activity increased over time by ERS and by applied IAA [G-3-PD(NAD)], (ii) activity increased over time by ERS but not by applied IAA (PFK, G-6-PD), (iii) activity increased over time but not by ERS or applied IAA (6-PGD). Increases in G-3-PD(NAD), G-6-PD, and PFK activity in leafy cuttings were positively related to primordium development. 6-PGD activity increased in leafy cuttings during primordium development and may have supported it. However, equal increases occurred in decapitated cuttings, in which the long-term development of primordia was supressed. Results for G-3-PD(NAD) that were obtained in an experiment with jack pine (Pinus banksiana Lamb.) seedling cuttings were similar to results for the same enzyme in bean cuttings. G-3-PD(NAD) activity in naphthaleneacetic acid-treated jack pine cuttings increased with time, in comparison with untreated cuttings, before root emergence.     

Adventitious root initiation in hypocotyl and epicotyl cuttings of eastern white pine (Pinus strobus) seedlings

The present paper reports results of experiments to develop a system for studying adventitious root initiation in cuttings derived from seedlings. Hypocotyl cuttings of 2-week-old eastern white pine (Pinus strobus L.) seedlings were treated for 5 min with 0, 100, 200, 300, 400, 500 or 600 mg l^?1 (0, 0.54, 1.07, 1.61, 2.15, 2.69 or 3.22 mM) 1-naphthaleneacetic acid (NAA) to determine the effect on root initiation. The number of root primordia per cutting was correlated with NAA concentration and the square of NAA concentration. Thus, the number increased from less than one per cutting in the 0 NAA treatment to approximately 40 per cutting at 300 mg l-1 NAA, above which no substantial further increase was observed. The larger number of root primordia formed in response to increasing concentrations of NAA was due to the formation of primordia over a larger proportion of the hypocotyls. Histological analysis of the timing of root primordium formation in hypocotyl cuttings revealed three discernible stages. Progression through these stages was relatively synchronous among NAA-treated hypocotyl cuttings and within a given cutting, but variation was observed in the portion of different cuttings undergoing root formation. Control-treated hypocotyl cuttings formed root primordia at lower frequencies and more slowly than NAA-treated cuttings, with fewer primordia per cutting. Epicotyl cuttings from 11-week-old seedlings also formed adventitious roots, but more slowly than hypocotyl cuttings. NAA treatment of epicotyl cuttings caused more rapid root initiation and also affected the origin of adventitious roots in comparison with nontreated cuttings. NAA-treated epicotyl cuttings formed roots in a manner analogous to that of the hypocotyl cuttings, directly from preformed vascular tissue, while control-treated epicotyl cuttings first formed a wound or callus tissue and subsequently differentiated root primordia within that tissue. This system of inducing adventitious roots in pine stem cuttings lends itself to studying the molecular and biochemical steps that occur during root initiation and development.     

Die Wirkung Des »Gibberellinantagonisten« 2-Chloräthyltrimethylammoniumchlorid (CCC) auf die Stecklingsbewurzelung Windender und nicht Windender Pflanzen

Summary Root initiation at cuttings is inhibited by application of gibberellin. CCC does not antagonize this inhibition. However, CCC stimulates root initiation at cuttings of three different species of twining plants known to be rich in endogenous gibberellin. CCC seems to be an antagonist for endogenous but not for exogenous gibberellin, which suggests that CCC influences gibberellin biosynthesis.     

Cytokinin Metabolism in Phaseolus vulgaris L.: I: VARIATIONS IN CYTOKININ LEVELS IN LEAVES OF DECAPITATED PLANTS IN RELATION TO LATERAL BUD OUTGROWTH

A stable isotope dilution method employing a deuterium-labelledinternal standard and combined gas chromatography-mass spectrometryhas been used to quantify the accumulation of di-hydrozeatin-O-ß-D-glucosidein the primary leaves of decapitated, disbudded bean plants.This cytokinin accumulated at a rate of 11 ng g^–1 fr.wt. d^–1 (eq. to an increase of 50 ng d^–1 per leaf),reaching a maximum of c. 500 ng g^–1 after 40 d from decapitation.This accumulation appeared to parallel the gradual increasein leaf fresh weight, and did not occur in detached leaves,in leaves of intact plants, or in leaves of plants that weredecapitated but not disbudded. When secondary lateral buds wereallowed to grow out from decapitated and initially disbuddedplants, the levels of dihydrozeatin-O-ß-D-glucosidein the primary leaves rapidly declined to a value similar toor lower than that found in leaves of intact plants. A similardecline in dihydrozeatin-O-ß-D-glucoside levels wasseen over 5 d in detached leaves of plants which had been decapitatedand disbudded for 15 d; this effect was reduced but not preventedwhen the leaves were supplied with inorganic nutrients. Theseresults are discussed in relation to the metabolism and distributionof cytokinins in the whole plant.     

Activity and isoforms of peroxidases, lignin and anatomy, during adventitious rooting in cuttings of Ebenus cretica L

Adventitious rooting of Ebenus cretica cuttings was studied in order to examine a) the rooting ability of different genotypes in relation to electrophoretic patterns of peroxidases. b) the activity and electrophoretic patterns of soluble and wall ionically bound peroxidases, the lignin content and anatomical changes in the control and IBA treated cuttings of and genotypes in the course of adventitious root formation. In addition, a fraction of soluble cationic peroxidases was separated by gel filtration chromatography from the total soluble peroxidases of a genotype. No rooting occurred in cuttings without IBA-treatment. In both genotypes, electrophoretic patterns of soluble anionic peroxidases revealed two common peroxidase isoforms, while a fast-migrating anionic peroxidase isoform (A3) appeared only in genotypes. Both genotypes showed similar patterns of soluble, as well as wall ionically bound cationic peroxidase isoforms. The number of isoforms was unchanged during the rooting process (induction, initiation and expression phase) but an increase in peroxidase activity (initiation phase) followed by decrease has been found in IBA-treated cuttings. During initiation phase the lignin content was almost similar to that on day 0 in genotype while it was reduced at by about 50% in genotype at the respective time. Microscopic observations revealed anatomical differences between genotypes. According to this study, the and genotypes display differences in anatomy, lignin content, activity of soluble peroxidases and the electrophoretic patterns of soluble anionic peroxidase isoforms. The A3-anionic peroxidase isoform could be used as biochemical marker to distinguish and genotypes of E. cretica and seems to be correlated to lignin synthesis in rooting process.     

Fluctuations of different endogenous phenolic compounds and cinnamic acid in the first days of the rooting process of cherry rootstock 'GiSelA 5' leafy cuttings

The relationship between the phenol composition of rooting zones and rootability was studied in the first days after the establishment of cuttings. The trial included two different types of cuttings (basal and terminal). Additionally, the influence of exogenously applied auxin (IBA) was observed. The best rooting results (55.6%) were achieved with terminal IBA treated cuttings, while only 1.9% of basal cuttings formed roots. The auxin treatment increased the root formation in terminal, but not in basal cuttings. Low rooting rate of basal cuttings was probably due to higher lignification rate of the basal tissue which can represent a mechanical barrier for root emergence. When measuring phenolic compounds and cinnamic acid, terminal cuttings contained higher (rutin, vanillic acid, (-)-epicatechin, caffeic acid and sinapinic acid) or equal concentrations of detected phenols as basal cuttings, while applied auxin did not influence the level of any of discussed phenolics, neither of cinnamic acid. It is to assume that cuttings for starting of root induction phase should contain certain levels of several phenolic compounds, but higher influence on rooting success is to be ascribed to the impact of the auxin level. During the time of the experiment concentrations of monophenols sinapinic acid and vanillic acid rapidly decreased. This decrease was more pronounced in terminal cuttings, which might have a better mechanism of lowering those two compounds to which a negative influence on rooting is ascribed. Fluctuations and differences between treatments of other phenolics were not significant enough to influence the rooting process.     

Effect of Auxins on Adventitious Root Development from Single Node Cuttings of Camellia sinensis (L.) Kuntze and Associated Biochemical Changes

An attempt was made to induce rooting from single node cuttings of Camellia sinensis var. TV-20 under controlled conditions and study its biochemical changes during rooting. The nodal cuttings were pretreated with different concentrations of IAA, NAA and IBA and kept in a growth chamber (25 ±2 °C, 16 h photoperiod (55 μ mol m⁻² s⁻¹) with cool, white fluorescent lamps and 65% relative humidity) for 12 h. Among the three auxins used for pretreatment, IBA showed more positive response on rooting as compared to IAA and NAA within 2 weeks of transfer to potting medium. Among four concentrations of IBA tested, 75 ppm gave maximum percentage of rooting, number of roots and root length. Therefore, IBA was used further in experiments for biochemical investigation. The adventitious rooting was obtained in three distinct phases i.e. induction (0–12 days), initiation (12–14 days) and expression (14–18 days). IAA-oxidase activity of IBA-treated cuttings increased slightly as compared to control. The activity was found to decrease during induction and initiation phases and increase during expression phase. The peroxidase activity in IBA-treated cuttings increased up to initiation phase and declined at the expression phase. Polyphenoloxidase activity increased both in IBA-treated and control cuttings during induction and initiation phase but declined slowly during expression phase. Total phenolic content was higher in IBA-treated cuttings, particularly in initiation and expression phases and it also correlated with peroxidase activity. Phenolics might be playing key role for induction of adventitious rooting, and phenolic compounds can be used as rooting enhancer in tea plant.     

Early steps of adventitious rooting: morphology,hormonal profiling and carbohydrate turnover in carnation stem cuttings

The rooting of stem cuttings is a common vegetative propagation practice in many ornamental species. A detailed analysis of the morphological changes occurring in the basal region of cultivated carnation cuttings during the early stages of adventitious rooting was carried out and the physiological modifications induced by exogenous auxin application were studied. To this end, the endogenous concentrations of five major classes of plant hormones [auxin, cytokinin (CK), abscisic acid, salicylic acid (SA) and jasmonic acid] and the ethylene precursor 1‐aminocyclopropane‐1‐carboxylic acid were analyzed at the base of stem cuttings and at different stages of adventitious root formation. We found that the stimulus triggering the initiation of adventitious root formation occurred during the first hours after their excision from the donor plant, due to the breakdown of the vascular continuum that induces auxin accumulation near the wounding. Although this stimulus was independent of exogenously applied auxin, it was observed that the auxin treatment accelerated cell division in the cambium and increased the sucrolytic activities at the base of the stem, both of which contributed to the establishment of the new root primordia at the stem base. Further, several genes involved in auxin transport were upregulated in the stem base either with or without auxin application, while endogenous CK and SA concentrations were specially affected by exogenous auxin application. Taken together our results indicate significant crosstalk between auxin levels, stress hormone homeostasis and sugar availability in the base of the stem cuttings in carnation during the initial steps of adventitious rooting.