Lateral root development and saponin accumulation as affected by IBA or NAA in adventitious root cultures of <Emphasis Type="Italic">Panax ginseng</Emphasis> C.A. Meyer

Summary The effect of indole-3-butyric acid (IBA) and 1-naphthaleneacetic acid (NAA) on lateral root formation was investigated in adventitious root culture of Panax ginseng. Lateral root formation was affected by IBA (24.6 μM) or NAA (9.8 μM). Lateral root primordia emerged from the explant root pericycle after about 7 d of culture when the roots were cultured on Schenk and Hildebrandt (SH) medium supplemented with 24.6 μM IBA or 9.8 μM NAA. However, no changes were observed in the explant root pericycle on auxin-free medium. The IBA treatment was more effective for lateral root induction and root growth compared to NAA. In morphological and histological aspects, the lateral roots formed under IBA treatment developed normally, while NAA-treated roots exhibited abnormal growth. The accumulation of total saponin was greater in roots treated with IBA than with NAA.     

Protein Changes Associated with Adventitious Root Formation in Hypocotyls of Pinus Radiata

The changes in soluble proteins associated with adventitious root formation in hypocotyls of radiata pine (Pinus radiata D. Don) were studied using one- and two-dimensional polyacrylamide gel electrophoresis. Protein content decreased during the first day after root excision, and kept decreasing till the end of the time course under non-rooting conditions, i.e., on medium without growth regulators, with indole-3-butyric acid (IBA) + kinetin, or with kinetic alone. During adventitious root initiation in response to IBA, however, the protein content began to increase from day 1 to its maximum at day 7, coinciding with the early stage of root initiation. A comparative analysis of protein changes by two-dimensional polyacrylamide gel electrophoresis showed 16 proteins that were probably associated with root initiation and development.     

Synergistic interaction between coumarin 1,2-benzopyrone and indole-3-butyric acid in stimulating adventitious root formation in Vigna radiata (L.) Wilczek cuttings: I. Endogenous free and conjugated IAA and basic isoperoxidases

Cuttings from 7-day-old Vigna radiata seedlings were treated for 24 h with various concentrations of coumarin and/or indole-3-butyric acid (IBA), applied either alone or in combination, in order to stimulate adventitious root formation (ARF). The effects of treatment on endogenous free and conjugated indole-3-acetic acid (IAA), basic peroxidase (basic PER) activity and its isoperoxidases analysis and their relation to ARF were then investigated at the potential rooting sites during the first 96 h after application. Simultaneously, combined treatments acted synergistically in inducing more adventitious roots in treated cuttings than in those treated with coumarin or IBA individually, as compared with the control. Endogenous free IAA increased transiently in treated cuttings as compared with the control and the maximum increase occurred with the combined treatment. This suggests that coumarin and IBA may act synergistically in increasing the endogenous free IAA level during the induction phase of rooting to initiate more roots. Likewise, higher level of conjugated IAA was also found in treated cuttings than in untreated ones, during the primary events of ARF, with the maximum level occurring in the combined treatment. Comparison of the dynamics of conjugated IAA and activity of basic PERs led to conclusion that the former but not the latter is responsible for downregulation of endogenous IAA levels significantly during the primary events of ARF. A sharp increases in basic PERs occurred during the secondary events of ARF, suggesting their role in root initiation and development rather than root induction.     

Influence of Aryl Esters of Indole-3-acetic and Indole-3-butyric Acids on Adventitious Root Primordium Initiation and Development

Synthetic aryl esters of indole-3-acetic acid (IAA) and indole-3-butyric acid (IBA) greatly enhanced adventitious root primordium initiation in bean (Phaseolus vulgaris L. cv. Top Crop) and jack pine (Pinus banksiana Lamb.) cuttings, respectively. Bean cuttings produced 95 to 154% more macroscopically visible root primordia in 2 days when treated with phenyl indole-3-acetate (P-IAA), in comparison with an equal concentration of IAA. Substantial but lesser increases occurred when treatment was done with 3-hydroxyphenyl indole-3-acetate (3HP-IAA). On a molar basis, either P-IAA or 3HP-IAA were 10 or more times as efficient as IAA in inducing adventitious root primordium initiation in bean cuttings. Methyl indole-3-acetate was no more effective than IAA in these tests. Phenyl indole-3-butyrate (P-IBA) consistently enhanced the number of rooted jack pine seedling cuttings by 11 to 12% in comparison with a 27% higher concentration of IBA. The number of elongated roots (2 mm or more) after 5 days was 165 to 276% greater for P-IAA than for IAA-treated bean cuttings. Similar but lesser increases occurred as a result of 3HP-IAA treatment. P-IBA in comparison with IBA treatment did not influence either the number of roots or length of the longest root per rooted jack pine cutting. Enzymes in bean and jack pine cuttings hydrolyzed the aryl esters. However, check experiments showed that initial integrity of the esters was required for enhanced activity in inducing root primordium initiation. Treatment of bean cuttings with hydrolysates of P-IAA, or with IAA and phenol, alone or combined, did not influence root primordium initiation or development in a manner different from treatment with IAA alone.     

Comparative effect of IBA, BSAA and 5,6-Cl2-IAA-Me on the rooting of hypocotyl in mung bean

Mung bean hypocotyl cuttings were treated with indole-3-butyric acid (IBA), 3-(benzo[b]selenienyl)acetic acid (BSAA) and 5,6-dichloroindole-3-acetic acid methyl ester (5,6-Cl2-IAA-Me) at different concentrations, respectively. Each chemical produced the maximum number of adventitious roots at a different concentration. Compared with IBA treatment, 5,6-Cl2-IAA-Me and BSAA treatments significantly increased root numbers on hypocotyl cuttings at lower concentration, particularly of 5,6-Cl2-IAA-Me treatment. Combinations of paclobutrazol (PB) with either 5,6-Cl2-IAA-Me or BSAA significantly stimulated the production of more adventitious roots than either chemical alone or combined. Capillary electrophoresis analysis have shown that the levels of IAA, IBA and BSAA in IBA plus PB or BSAA plus PB treatments were higher than those of IBA or BSAA alone. It was suggested that the cause of the synergistic effect of IBA (or BSAA) plus PB treatment might be due to increased endogenous auxin level. The activities of peroxidase and IAA oxidase in the rooting zone coincided with root development, indicating that the activities of these two enzymes were positively correlated to rooting. Peroxidase and IAA oxidase activity in all treatments started 24 h and 12 h after cutting, respectively. It is suggested that the major role of IAA oxidase differed from that of peroxidase in adventitious root formation.     

Comparison of movement and metabolism of indole-3-acetic acid and indole-3-butyric acid in mung bean cuttings

Indole-3-butyric acid (IBA) was much more effective than indole-3-acetic acid (IAA) in inducing adventitious root formation in mung bean ( Vigna radiata L.) cuttings. Prolonging the duration of treatment with both auxins from 24 to 96 h significantly increased the number of roots formed. Labelled IAA and IBA applied to the basal cut surface of the cuttings were transported acropetally. With both auxins, most radioactivity was detected in the hypocotyl, where roots were formed, but relatively more IBA was found in the upper sections of the cuttings. The rate of metabolism of IAA and IBA in these cuttings was similar. Both auxins were metabolized very rapidly and 24 h after application only a small fraction of the radioactivity corresponded to the free auxins. Hydrolysis with 7 M NaOH indicates that conjugation is the major pathway of IAA and IBA metabolism in mung bean tissues. The major conjugate of IAA was identified tentatively as indole-3-acetylaspartic acid, whereas IBA formed at least two major conjugates. The data indicate that the higher root-promoting activity of IBA was not due to a different transport pattern and/or a different rate of conjugation. It is suggested that the IBA conjugates may be a better source of free auxin than those of IAA and this may explain the higher activity of IBA.     

Endogenous indole-3-acetic acid during adventitious root formation inPopulus £canadensis Moench.

Indole-3-butyric acid (IBA), phenylacetic acid (PAA) and naphthaleneacetic acid (NAA) were applied at a concentration of 10^-4 mol dm^-3 to stem cutting bases ofPopulus x canadensis Moench. During adventitious root formation, the content of indole-3-acetic acid (IAA) in cutting bases was estimated using the fluorimetric method. In the control variant, a rapid increase in endogenous IAA appeared after 24-h cultivation followed by gradual decrease during the following days. In contrast, the variants treated with IBA, PAA, and especially NAA exhibited firstly a decrease in endogenous IAA content and only afterwards an increase, reaching a maximum 48 h after excision. As root regeneration proceeded gradually, a decrease in the level of endogenous IAA occurred in all treatments. The first adventitious roots appeared in all treatments after 216-h cultivation.     

Effect of Exogenous Indole-3-Acetic Acid and Indole-3-Butyric Acid on Internal Levels of the Respective Auxins and Their Conjugation with Aspartic Acid during Adventitious Root Formation in Pea Cuttings

The influence of exogenous indole-3-acetic acid (IAA) and indole-3-butyric acid (IBA) on the internal levels of these auxins was studied during the first 4 days of adventitious root formation in cuttings of Pisum sativum L. The quantitations were done by high performance liquid chromatography with spectrofluorometric detection. IBA, identified by combined gas chromatography-mass spectrometry (GC-MS), was found to naturally occur in this plant material. The root inducing ability of exogenous IBA was superior to that of IAA. The IAA level in the tissue increased considerably on the first day after application of IAA, but rapidly decreased again, returning to a level twice the control by day 3. The predominant metabolic route was conjugation with aspartic acid, as reflected by the increase in the level of indole-3-acetylaspartic acid. The IBA treatment resulted in increases in the levels of IBA, IAA, and indole-3-acetylaspartic acid. The IAA content rapidly returned to control levels, whereas the IBA level remained high throughout the experimental period. High amounts of indole-3-butyrylaspartic acid were found in the tissue after feeding with IBA. The identity of the conjugate was confirmed by ¹H-nuclear magnetic resonance and GC-MS. IBA was much more stable in solution than IAA. No IAA was detected after 48 hours, whereas 70% IBA was still recovered after this time. The relatively higher root inducing ability of IBA is ascribed to the fact that its level remained elevated longer than that of IAA, even though IBA was metabolized in the tissue. Adventitious root formation is discussed on the basis of these findings.     

Auxins and polyamines in relation to differential in vitro root induction on microcuttings of two pear cultivars

The internal levels of indole-3-acetic acid (IAA) and polyamines (PAs) and the metabolism of indole-3-butyric acid (IBA) were studied in relation to the in vitro rooting process of two pear cultivars, the easy-to-root Conference and the difficult-to-root Doyenne d'Hiver. Doyenne d'Hiver required about a 10 times higher concentration of IBA to achieve a rooting percentage similar to that of Conference. One- or two-day exposures to IBA were sufficient to stimulate rooting but with different efficiency for each cultivar. Longer exposure to auxin strongly increased the root number in Conference, whereas root elongation was inhibited in both cultivars. The metabolism of IBA in both cultivars was not significantly different when IBA was used at a high concentration to stimulate maximal rooting in Doyenne d'Hiver. IBA was mainly conjugated into IBA glucose, which was accumulated, and a small amount was converted into free IAA in both cultivars. However, in Doyenne d'Hiver this metabolic pathway appears to be active only at a higher exogenous IBA concentration. At a high IBA concentration more callus was formed by Doyenne d'Hiver, indicating that the cells of Doyenne d'Hiver are not capable of responding to the hormone in the same manner as Conference cells. Anatomic observations indicated that the capacity to induce initial dividing cells was more efficient in Doyenne d'Hiver, but subsequently the number of root primordia formed and root development were much reduced relative to Conference. A possible correlation between these processes and an early increase followed by a decrease of free IAA was seen in Conference. By day 4, a significant increase in IAA conjugates and free putrescine was observed in Doyenne d'Hiver. This higher putrescine content may be related to the lower amount of root development. Together with previous studies these results indicate that differences in the uptake and metabolism of applied auxins may affect rooting ability and the subsequent development of adventitious roots in microcuttings of pear.Abbreviations IBA indole-3-butyric acid - IAA indole-3-acetic acid - PA(s) polyamine(s) - HPLC high pressure liquid chromatography - GC-MS gas chromatography-mass spectrometry - TCA trichloroacetic acid dansyl, 1-dimethylaminonaphthalene-5-sulfonyl - TLC thin layer chromatography - TBA terbutilic alcohol - IBAGluc IBA glucose - IAAGluc IAA glucose - IAAsp IAA aspartate     

The induction of contractile roots in Gladiolus grandiflorus

The number of contractile roots formed in gladioli was inversely related to the depth of the planted corm. Below a certain depth, no contractile roots were produced. Large corms did not produce contractile roots at any planting depth but produced two or more small corms. Depth perception is a function of two independent mechanisms, namely, temperature fluctuations in the root-initiation zone and the amount of light perceived mainly by the upper sheath leaf, the length of which varies with planting depth. Various growth substances applied to the leaves or corms did not induce contractile roots in dark-grown plants but roots were induced by indole-3-butyric acid in both small and large corms grown at constant temperatures and light. Abscisic acid retarded the formation of contractile roots under inductive conditions.Abbreviations CR contractile root(s) - IBA indole-3-butyric acid     

Characteristics of adventitious root formation in cotyledon segments of mango (Mangifera indica L. cv. Zihua): two induction patterns, histological origins and the relationship with polar auxin transport

Cotyledon segments derived from zygote embryos of mango (Mangifera indica L. cv. Zihua) were cultured on agar medium for 28 days. Depending on different pre-treatments with plant growth regulators, two distinct patterns of adventitious roots were observed. A first pattern of adventitious roots was seen at the proximal cut surface, whereas no roots were formed on the opposite, distal cut surface. The rooting ability depended on the segment length and was significantly promoted by pre-treatment of embryos with indol-3-acetic acid (IAA) or indole-3-butyric acid (IBA) for 1 h. A pre-treatment with the auxin transport inhibitor 2,3,5-triiodobenzoic acid (TIBA) completely inhibited adventitious root formation on proximal cut surfaces. A second pattern of roots was observed on abaxial surfaces of cotyledon segments when embryos were pre-treated with 2,700 μM 1-naphthalenacetic acid (NAA) for 1 h. Histological observations indicated that both patterns of adventitious roots originated from parenchymal cells, but developmental directions of the root primordia were different. A polar auxin transport assay was used to demonstrate transport of [³H] indole-3-acetic acid (IAA) in cotyledon segments from the distal to the proximal cut surface. In conclusion, we suggest that polar auxin transport plays a role in adventitious root formation at the proximal cut surface, whereas NAA levels (influx by diffusion; carrier mediated efflux) seem to control development of adventitious roots on the abaxial surface of cotyledon segments.     

Adventitious rooting performance in micropropagated Cornus mas

Axillary buds sampled from a mature 27-year-old Cornus mas cv. Macrocarpa were grown in vitro on modified woody plant medium (WPM). Adventitious rooting performance of microshoots was assayed on half-strength WPM supplemented with 1-naphthaleneacetic acid (NAA) or indole-3-butyric acid (IBA) under various pH. NAA induced significantly higher rooting frequencies than IBA. The pH of 6.8 inhibited rooting, and differentiated roots were extremely thick and fragile. The highest rooting frequency was recorded on half-strength WPM supplemented with 5.37 µM NAA at the pH value adjusted to 6.2 (73 % of rooted shoots). In the presence of IBA, the formation of adventitious roots was observed only in the basal part of the microshoot dipped into rooting medium. In the case of NAA, however, adventitious roots arose also from the parts of microshoots that were not in contact with medium. The growth of aerial roots was always positively gravitropic. The nuclear microsatellite Cf-G17 gave a monomorphic fingerprinting pattern across the mother shrub and micropropagated plantlets. Acclimatized plants did not show any visually detectable morphological variation and the aerial adventitious root formation was no longer observed.     

Relationship of indole-3-butyric acid and adventitious rooting in M.26 apple (Malus Pumila mill.) shoots cultured in vitro

The role of indole-3-butyric acid (IBA) in adventitious root formation was studied by analyzing the uptake and subsequent metabolism of IBA in shoots of M.26 apple (Malus pumila Mill.) rootstock grown in vitro. Roots were induced by exposing shoots to 4 M IBA and [³H]IBA for 5 days in the dark and then transferring them to plant growth regulator (PGR)-free medium in the light until roots formed. Approximately 50% of the total radioactivity applied was taken up from the agar medium by the shoots during the 5-day incubation period in IBA. Indole-3-butyric acid metabolism was studied by extraction and high-performance liquid chromatographic (HPLC) separation of [³H]IBA and metabolites from the basal sections of treated shoots. The major [³H]IBA metabolite co-eluted with authentic [¹⁴C]indole-3-acetic acid (IAA) suggesting that IBA was converted to IAA in the shoots. The proportion of newly synthesized IAA present as conjugates was higher at the end of the 5-day IBA treatment period than after 13 days in PGR-free medium. There appeared to be no conjugation of IBA at any time.     

植物生长调节剂对鹿角杜鹃扦插繁殖的影响

以腐殖土为基质,研究了5种植物生长调节剂的4个浓度处理、以及萘乙酸(NAA)不同浓度与浸泡时间处理对鹿角杜鹃(Rhododendron latoucheae)当年生枝条扦插繁殖12个育苗指标的影响及各指标间的相关性,并运用隶属函数法对各处理组合的育苗效果进行了综合评价。结果表明:5种生长调节剂中,赤霉素(GA3)、吲哚丁酸(IBA)处理在大部分育苗指标上表现良好,为鹿角杜鹃扦插育苗的理想调节剂,吲哚乙酸(IAA)效果次之,6-苄氨基嘌呤(6-BA)、NAA处理的育苗指标表现较差;4个浓度水平上,愈伤率、腐烂率及老叶留存率总体以低浓度处理最佳,随着浓度升高效果下降;其它9个指标对浓度的响应因生长调节剂种类而异。各指标相关性分析发现,4个根系生长指标,即不定根数、最长不定根长、总根数及根系直径之间呈极显著正相关,但它们与愈伤率、腐烂率、生根率及老叶留存率、新梢率间的相关性较弱。隶属函数综合评价表明,50 mg/L GA3处理为鹿角杜鹃扦插育苗的最佳组合,其次为100 mg/L GA3和400 mg/L IBA处理。本研究为杜鹃花产业化育苗提供了科学依据。     

间苯二酚、水杨酸对绿豆下胚轴不定根形成的作用

２０—１００ｍｇＬ（－１）间苯二酚能明显地促进绿豆下胚轴不定根的形成,与２０ｍｇＬ（－１）ＩＢＡ混合处理具加成效应,其作用在于降低生根初期ＩＡＡ氧化酶和多酚氧化酶活性．１０—１００ｍｇＬ（－１）水杨酸抑制下胚轴不定根的形成,随处理浓度的加大,对生根数目、生根范围和根重的抑制作用增加．水杨酸处理后１－３ｄ,能提高ＩＡＡ氧化酶和多酚氧化酶的活性．     

Differential responses of primary and lateral roots to indole-3-acetic acid,indole-3-butyric acid,and 1-naphthaleneacetic acid in maize seedlings

The role of auxins on root system architecture was studied by applying indole-3-acetic acid (IAA), indole-3-butyric acid (IBA), and 1-naphthaleneacetic acid (NAA) to maize roots and analysing the main processes involved in root development: primary root (PR) elongation, lateral root (LR) formation, and LR root elongation. We found that these effects were not dependent only on concentration, but also on the type of auxin applied. We also studied temporal changes in auxin inhibition of PR elongation. These temporal changes were analysed calculating the elongation ratio between two consecutive one day periods after auxin application. It was observed that a reduction in root elongation was also dependent on the type of auxin applied and its concentration. The inhibitory effect of IBA and IAA decreased on the second day, and the ratio also increased with the concentration. In contrast, NAA increased root elongation inhibition with time. Indeed, the ratio decreased as the NAA concentration increased. Regarding LR formation, we observed that external auxin increased only LR formation in certain zones of the PR. Finally, comparison of inhibition elongation associated with auxin in the LR and PR clearly demonstrates that PR elongation was more sensitive to auxin than LR elongation.     

Ethylene and auxin-ethylene interaction in adventitious root formation in mung bean (Vigna radiata) cuttings

The role of ethylene in adventitious root formation and its involvement in auxin-induced rooting were investigated in cuttings ofVigna radiata (L.). Treatment with 30 M indole-3-acetic acid (IAA) for 24 h slightly inhibited rooting, whereas the same concentration of indole-3-butyric acid (IBA) significantly stimulated it. Ethylene derived from 1-aminocyclopropane-1-carboxylic acid (ACC) increased the number of adventitious roots but inhibited their emergence and elongation. Endogenous levels of ethylene, ACC, and malonyl-ACC (MACC) were initially higher in cuttings treated with IAA. This trend was quickly reversed, and cuttings, particularly hypocotyls, treated with IBA produced higher levels of ethylene and had more ACC and MACC during most of the rooting process. Aminoethoxyvinylglycine significantly inhibited rooting, but its inhibitory effect could not be reversed by ACC. The data suggest that the stimulating effect of IBA on rooting is closely associated with its induction of ACC and ethylene biosynthesis.     

Root Formation in Ethylene-Insensitive Plants

Experiments with ethylene-insensitive tomato (Lycopersicon esculentum) and petunia (Petunia x hybrida) plants were conducted to determine if normal or adventitious root formation is affected by ethylene insensitivity. Ethylene-insensitive Never ripe (NR) tomato plants produced more below-ground root mass but fewer above-ground adventitious roots than wild-type Pearson plants. Applied auxin (indole-3-butyric acid) increased adventitious root formation on vegetative stem cuttings of wild-type plants but had little or no effect on rooting of NR plants. Reduced adventitious root formation was also observed in ethylene-insensitive transgenic petunia plants. Applied 1-aminocyclopropane-1-carboxylic acid increased adventitious root formation on vegetative stem cuttings from NR and wild-type plants, but NR cuttings produced fewer adventitious roots than wild-type cuttings. These data suggest that the promotive effect of auxin on adventitious rooting is influenced by ethylene responsiveness. Seedling root growth of tomato in response to mechanical impedance was also influenced by ethylene sensitivity. Ninety-six percent of wild-type seedlings germinated and grown on sand for 7 d grew normal roots into the medium, whereas 47% of NR seedlings displayed elongated tap-roots, shortened hypocotyls, and did not penetrate the medium. These data indicate that ethylene has a critical role in various responses of roots to environmental stimuli.