Indole-3-butyric acid in plants: occurrence, synthesis, metabolism and transport

Indole-3-butyric acid (IBA) was recently identified by GC/MS analysis as an endogenous constituent of various plants. Plant tissues contained 9 ng g^?1 fresh weight of free IBA and 37 ng g^?1 fresh weight of total IBA, compared to 26 ng g^?1 and 52 ng g^?1 fresh weight of free and total indole-3-acetic acid (IAA), respectively. IBA level was found to increase during plant development, but never reached the level of IAA. It is generally assumed that the greater ability of IBA as compared with IAA to promote rooting is due to its relatively higher stability. Indeed, the concentrations of IAA and IBA in autoclaved medium were reduced by 40% and 20%, respectively, compared with filter sterilized controls. In liquid medium, IAA was more sensitive than IBA to non-biological degradation. However, in all plant tissues tested, both auxins were found to be metabolized rapidly and conjugated at the same rate with amino acids or sugar. Studies of auxin transport showed that IAA was transported faster than IBA. The velocities of some of the auxins tested were 7. 5 mm h^?1 for IAA, 6. 7 mm h^?1 for naphthaleneacetic acid (NAA) and only 3. 2 mm h^?1 for IBA. Like IAA, IBA was transported predominantly in a basipetal direction (polar transport). After application of ³H-IBA to cuttings of various plants, most of the label remained in the bases of the cuttings. Easy-to-root cultivars were found to absorb more of the auxin and transport more of it to the leaves. It has been postulated that easy-to-root, as opposed to the difficult-to-root cultivars, have the ability to hydrolyze auxin conjugates at the appropriate time to release free auxin which may promote root initiation. This theory is supported by reports on increased levels of free auxin in the bases of cuttings prior to rooting. The auxin conjugate probably acts as a ‘slow-release’ hormone in the tissues. Easy-to-root cultivars were also able to convert IBA to IAA which accumulated in the cutting bases prior to rooting. IAA conjugates, but not IBA conjugates, were subject to oxidation, and thus deactivation. The efficiency of the two auxins in root induction therefore seems to depend on the stability of their conjugates. The higher rooting promotion of IBA was also ascribed to the fact that its level remained elevated longer than that of IAA, even though IBA was metabolized in the tissue. IAA was converted to IBA by seedlings of corn and Arabidopsis. The K_m value for IBA formation was low (approximately 20 μM), indicating high affinity for the substrate. That means that small amounts of IAA (only a fraction of the total IAA in the plant tissues) can be converted to IBA. It was suggested that IBA is formed by the acetylation of IAA with acetyl-CoA in the carboxyl position via a biosynthetic pathway analogous to the primary steps of fatty acid biosynthesis, where acetyl moieties are transferred to an acceptor molecule. Incubation of the soluble enzyme fraction from Arabidopsis with ³H-IBA, IBA and UDP-glucose resulted in a product that was identified tentatively as IBA glucose (IBGIc). IBGIc was detected only during the first 30 min of incubation, showing that it might be converted rapidly to another conjugate.     

Auxin Synergists in Rooting of Cuttings

Leafy cuttings of Eranthemum tricolor were treated with tannic acid, gallic acid, p-hydroxybenzoic acid and salicylic acid at the concentrations of 1000, 100, 10 and 1 nig/1 for 24 hours, whereafter they were dipped quickly in a 1000 mg/l solution of IAA, IBA and NAA for ten seconds. None of the phenolics showed any root promoting effect when used singly. In combination with NAA and IBA tannic acid promoted rooting, however, with IAA there was no effect to be seen. Gallic acid also markedly increased the number of roots of cuttings treated with NAA and IBA. Even in this case there was no effect with IAA. Synergism was also recorded between p-hydroxybenzoic acid and IAA or NAA but not with IBA. Salicylic acid greatly promoted rooting in combination with both IAA, IBA and NAA.     

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.     

Transport and metabolism of indole-3-butyric acid in cuttings of Leucadendron discolor

Indole-3-butyric acid (IBA) greatly enhanced the rooting of an early-flowering variety of protea, Leucadendron discolor, but had very little effect on a late-flowering variety. IBA transport and metabolism were studied in both varieties after incubating the cuttings in ³H-IBA. More of the radio-label was transported to the leaves of the easy-to-root variety than the difficult-to-root (35–45% and 10%, respectively). IBA was metabolized rapidly by the cuttings of both varieties and after 24 h most of the label was in the new metabolite. However, free IBA (about 10%) was present in the cuttings during the whole period up to the time of root emergence (4 weeks). More free IBA was accumulated in the base of easy-to-root cuttings, while in the difficult-to-root variety most of the IBA was found in the leaves. The metabolite was identified tentatively as an ester conjugate with a glucose. It is possible that IBA-glucose serves as a source for free IBA, and the difference between the varieties is a consequence of the free IBA which is released, transported and accumulated in the site of a root formation.     

Vegetative growth retardation,improved rooting and viability of olive cuttings in response to application of growth retardants

Spray and soil treatments of paclobutrazol and uniconazole were applied to young and mature olive plants and olive cuttings. Two clear phases, were found in the growth response of olive shoots to growth retardants: an early phase, which retards and even inhibits growth considerably; and a later phase, during which the shoots are released from the retardation and start to elongate rapidly. A somewhat slower response of the plants to soil application than to spray application of growth retardants was noticed. Paclobutrazol enhanced the rooting of cv. Manzanillo cuttings, whether applied to the mother plants or to the cuttings themselves. Indole-3-butyric acid (IBA) was needed in both cases. Sprouting was shown to reduce rooting. Paclobutrazol significantly inhibited sprouting and increased the meristematic activity in the base of the cuttings. Rooting of the hard-to-root cv. Kalamata was not enhanced by the treatments although callus formation was induced and viability was prolonged.     

植物生长调节剂对几种园林植物插条生根的影响

 我们用NAA、IBA、IAA等植物生长调节剂促进多种园林植物插条发根的试验。结果表明：菊花、大丽花插枝,经0.1mg/L和0.2 mg/L NAA处理,具有促进发根的明显效果;0.1mg／L和0.2 mg/L的NAA和IBA处理较难发根的印度黄檀插枝,其发根率则有显著提高;月季、大叶黄杨、水杉、池杉、海南石梓等植物插枝,虽都具有较高的发根率,但经不同浓度NAA和IBA浸渍处理后,其发根率仍有不同程度的提高。     

Metabolic changes during rooting in stem cuttings of five mangrove species

Vegetative propagation through rooting in stem cuttings in five tree mangroves namely Bruguiera parviflora, Cynometra iripa, Excoecaria agallocha, Heritiera fomes, and Thespesia populnea using IAA, IBA and NAA was reported. Spectacular increase in the root number was noted in the cuttings of H. fomes and C. iripa treated together with IBA (5000 ppm) and NAA (2500 ppm). The highest number of roots was obtained with IBA (2500 ppm) and NAA (500 ppm) in E. agallocha. B. parviflora and T. populnea responded better to IAA and IBA treatment. The species specific variation in the rooting response to exogenous application of auxins was reflected in the metabolic changes during initiation and development of roots in cuttings. Biochemical analysis showed increase of reducing sugar in the above-girdled tissues at initiation as well as subsequent development of roots which was further enhanced by the use of auxins. Decreases in the total sugar, total carbohydrate and polyphenols and increase in total nitrogen were recorded in the girdled tissues and the high C/N ratio at the initial stage helped in initiation of roots in all the species. Interaction of IBA and NAA promoted starch hydrolysis better than IAA and IBA during root development and subsequently reduced the C/N ratio and increased the protein-nitrogen activity during root development which suggest the auxin influenced mobilization of nitrogen to the rooting zone.Abbreviations IAA Indole-3-acetic acid - IBA Indole-butyric acid - NAA A-naphthalene acetic acid     

吲哚丁酸处理桉树插条后过氧化物酶活性和同工酶变化与插条生根的关系

尾叶桉MLA无性系（简称MLA）为难生根植物,属叶桉U6无性系（简称6）和刚果12号桉W5无性系（简称W5）为相对易生根植物。MLA的插条中的过氧化物酶（POD）活性较U6、W5的高。用吲哚丁酸（IBA）处理桉树的插条后,在扦插生根的不同阶段,插条内的POD活性呈现规律性的变化。蛋白质含量呈上升趋势。POD同工酶谱带也随生根的进程出现增多的现象。本文讨论了过氧化物酶与桉树插条生根的关系。     

Influence of IBA and Cordycepin on Rooting and RNA Synthesis in Stem Cuttings of Phaseolus aureus Roxb

Adventitious roots are initiated on stem cuttings of Phaseolusaureus Roxb. by treatment with IBA for 24 h, although subsequenttransfer to boric acid is essential for their development. Cordycepinenhances auxin-induced rooting when supplied for 4 h withinthe first twenty hours of IBA treatment, but not thereafter.Cordycepin alone does not enhance rooting. IBA treatment ofcuttings for 12 h results in a marked inhibition of RNA synthesis,including poly(A)-rich RNA, in the hypocotyl. After 24 h treatmentRNA synthesis is seen to increase, with a more marked recoveryin the synthesis of poly(A)+RNA relative to other RNAs. Subsequenttransfer to boric acid maintains this recovery. Cordycepin doesnot inhibit RNA synthesis below the level induced by IBA althoughon subsequent transfer to boric acid is seen to enhance synthesisand turnover of both polyadenylated and non-polyadenylated RNA. (Received August 28, 1982; Accepted November 25, 1982)     

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.     

Effect of Auxins and Light on Rooting Stem Cuttings of Populus nigra Salix tetrasperma, Ipomea fistulosa and Hibiscus notodus in Relation to Polarity

The apical and basal ends of stem cuttings of Populus nigra, Salix tetrasperma, Ipomoea fistulosa and Hibiscus notodus were treated with 10 mg/l solutions of IAA and IBA for 24 hours and were planted either erect or inverted both in light and dark. Observations for the number of cuttings that rooted and the roots produced on them were recorded at weekly intervals. In Salix, Ipomoea and Hibiscus rooting was more on cuttings planted erect, while in populus it did not differ much with the manner of planting. The reduced rooting in inverted cuttings may be ascribed to the low level of endogenous auxin at the apex due to polar transport. An exogenous application of auxins enhanced rooting on inverted cuttings. In dark, roots on Populus and Salix cuttings were produced both above and within the rooting medium. The weak polarity of these two plants may be due to the potential root primordia reported in their stem. The formation of callus occurred on the top of Populus cuttings whether planted erect or inverted but it differentiated into branches on erect cuttings only. In those planted in an inverted position the callus failed to differentiate in spite of the application of kinetin, auxins, TIBA, coumarin and sucrose, and dried ultimately.     

Effect of abscisic acid on endogenous IAA, auxin protector levels and peroxidase activity during adventitious root initiation in Vigna radiata cuttings

Endogenous levels of free and conjugated IAA, auxin protectors (Prs) and peroxidase (PER) activity and their relation to adventitious root initiation (ARI) were investigated at the potential sites of adventitious rooting in relation to exogenous application of 250 μM ABA during the first 120 h after treatment. Cuttings from 7-day-old mung bean [Vigna radiata (L.) Wilcz.] seedlings were treated with 125, 250, and 500 μM ABA for 24 h. ABA significantly stimulated ARI but extremely inhibited epicotyl growth as compared to control. Free and conjugated IAA were measured by reversed-phase high performance liquid chromatography while Prs and PER activities were measured spectrophotometrically. The present results also indicate that endogenous free IAA levels peaked later in ABA-treated cuttings than that in control, suggesting that ABA extended the length of the induction phase of rooting process in treated cuttings and that might explain the significant delay of the appearance of roots at the treated cuttings. Higher level of IAA conjugates was found in ABA-treated cuttings than that in untreated ones. Pr level also peaked later in ABA-treated cuttings than that in control, indicating that ABA extended the period of Pr activity. An initial temporary decrease of PER activity was found in associating with high levels of free IAA and Prs during most of the primary events, while the opposite occurred during the secondary events of adventitious rooting process in both treated and untreated cuttings. Thus, ABA may stimulate ARI in mung bean Vigna radiata cuttings by regulating the concentration and /or activities of endogenous IAA, Prs, and PER activity in favor of inducing a large number of adventitious roots at their potential sites of adventitious rooting.     

Effect of Ethephon, Indole Butyric Acid, and Treatment Solution pH on Rooting and on Ethylene Levels within Mung Bean Cuttings

Light-grown mung bean (Phaseolus aureus Roxb.) cuttings were treated with buffered and nonbuffered solutions of Ethephon, indole butyric acid (IBA), and the combination of both. Ethephon treatment resulted in increased tissue ethylene levels with increasing solution pH, but had no effect on rooting. IBA treatment had no effect on tissue ethylene levels, but strongly promoted rooting. Combinations of Ethephon and IBA had no effect on rooting of mung bean cuttings beyond that obtained by IBA alone.     

不同株龄日本落叶松插穗的内源激素含量与生根的关系

研究了日本落叶松母株年龄、插穗内源激素含量与生根之间的关系,以及外源IBA对插穗内源激素含量的影响及其对插穗生根的促进作用。结果表明：不同株龄插穗生根性状及插穗茎和叶中激素含量差异均达极显著水平,叶中激素含量对插穗生根力没有直接影响;插穗茎中生根抑制激素（ABA）含量随株龄增长而增加,生根促进激素与抑制激素的比值（IAA＋GA＋ZR）/ABA却随株龄的增长而递减,与生根力随株龄的变化趋势一致,且该比值与生根性状紧密相关,因此可作为评价母株（无性系）生根力的指标;插后13～32d是插穗愈伤组织形成和不定根诱导的关键期,此期生根促进激素消耗量大,茎中含量大幅度降低,进入根伸长生长阶段,含量上升;外源IBA促进插穗生根的机制在于通过外源激素的刺激,在不定根诱导期,插穗茎中ABA含量大幅度降低,从而有利于不定根的发生和发育。     

渐狭叶烟草组培苗生根条件优化及在基因转化中的应用

针对渐狭叶烟草(Nicotiana attenuata)组培苗生根困难的问题，本研究测试了不同浓度萘乙酸(NAA)、吲哚丁酸(IBA)、吲哚乙酸(IAA)、嘧啶醇、矮壮素、生根粉及活性炭等外源添加物对组培苗生根的影响。结果表明，对照处理的组培苗在90 d内基本不生根；不同浓度矮壮素处理也不能诱导组培苗生根；而合适浓度的NAA、IBA或IAA处理均能较好地诱导组培苗生根，其中0.8 mg·L^-1 IBA不仅具有较好的诱导生根效果，生根率达72.73%，且根及地上部分均生长良好。进一步测试发现0.8 mg·L^-1 IBA处理对转基因组培苗具有良好的生根效果，20 d后生根率达60%以上，而对照组未生根；经伤口诱导和0.8 mg·L^-1 IBA处理，40 d内总生根率可达90%以上。该研究为高效获得渐狭叶烟草转基因生根苗提供了解决方案，也为渐狭叶烟草作为烟草属模式植物的研究提供了便利。     

N-phenyl indolyl-3-biityraitiide and phenyl indole-3-thiolobutyrate enhance adventitious root primordium development

N-phenyl indoryl-3-butyramide (NP-IBA) and phenyl indole-3-thiolobutyrate (P-ITB) are new synthetic auxins that enhance rooting of bean ( Phaseolus vulgaris cv. Top Crop) hypocotyl cuttings and jack pine ( Pinus banksiana Lamb.) seedling cuttings. In comparison with an equal concentration of indole-3-butyric acid (IBA), NP-IBA treatment resulted in greater rooting of bean and jack pine seedling cuttings. IBA and P-ITB treatment at equal concentration were equally effective in inducing rooting of bean cuttings. However, in comparison with an equal concentration of IBA, P-ITB treatment promoted greater rooting of jack pine seedling cuttings, but P-ITB treatment was less effective than NP-IBA treatment. Application of crystalline chemical to jack pine seedling cuttings indicated lesser toxic effects and earlier and greater root-promoting effects of P-ITB and NP-IBA treatments, in comparison with IBA treatment. The earlier rooting of NP-IBA-treated jack pine seedling cuttings was positively related to increase in fresh weight of basal 1 cm stem before primordia were initiated. Weight of basal steins of NP-IBA-treated cuttings initially increased during propagation at day 2 in comparison with the control, and at day 3 in comparison with IBA-treated cuttings. Results that were obtained with seedling jack pine cuttings were generally confirmed for cuttings from 7-year-old trees.     

沙生柽柳扦插生根过程插穗相关理化特征分析

选取沙生柽柳半木质化枝条进行苗床扦插,通过实验测定插穗生根过程中内源激素(IAA、GA3、ZR、ABA)含量、可溶性营养物质(糖、蛋白质)含量及相关氧化酶(PPO、POD、SOD、IAAO)活性的动态变化特征,探讨沙生柽柳插穗扦插生根机理。结果表明:(1)沙生柽柳插穗内源激素含量随生根进程而发生变化,其中,IAA含量在扦插35d最大,并出现较大的波动变化;ZR含量在扦插55d前后变化明显,呈现低水平向高水平转化趋势;ABA、GA3含量依次呈先升高后降低再升高的变化过程,并在扦插15d和55d(80d)呈现变化的峰值和谷值。(2)沙生柽柳扦插生根与相关氧化酶活性密切相关,其中,POD、IAAO活性在插穗扦插35d后长时间保持较高水平,直至插穗生根后POD活性明显降低,IAAO活性有所增加;PPO、SOD活性则在插穗扦插15d保持较高活性,且PPO活性的变化均匀,SOD活性的高低交替变化明显。(3)在沙生柽柳扦插生根期间,插穗可溶性糖含量呈现生根前消耗减少与生根后积累增加两大变化过程,可溶性蛋白质含量表现为扦插后逐步积累增加的变化趋势。研究表明,高水平的IAA、ZR和低水平的GA3、ABA共同调控着沙生柽柳插穗生根;IAA能够通过促进插穗POD、PPO、IAAO活性变化来影响生根,较高的POD、IAAO活性可调节插穗IAA水平,高水平的PPO活性则催化插穗IAA-酚酸复合物的形成,进而诱导插穗生根。     

Auxin stability and accumulation during in vitro shoot morphogenesis influences subsequent root induction and development in <Emphasis Type="Italic">Eucalyptus grandis</Emphasis>

Recent results showed that after 16 months in the field, micropropagated eucalyptus plants have an inferior root system to cuttings. Such differences may be due to the plant growth regulators supplied during the culture stages of standard protocols, which are targeted at optimising plantlet yields and not root quality. This study investigated such a proposal, focusing on auxins in an easy-to-root clone. Initial results showed that the auxin provided in the standard protocol (NAA for multiplication and IBA for elongation) enabled 100% rooting in auxin-free medium, where rooting was faster than on IBA-rooting media. When auxin supply was omitted from multiplication and restricted to NAA or IAA during elongation, rooting in an auxin-free medium was reduced to 68 and 31%, respectively, reflecting the stabilities of these auxins in plant tissues. Additionally, 15% of shoots from the NAA-medium and 65% from the IAA-medium produced roots with altered graviperception. GC–MS analysis of these shoots revealed a relationship between free IAA-availability and altered graviperception. This was further tested by adding the IAA-specific transport inhibitor 2,3,5-triiodobenzoic acid to rooting media with IBA, IAA or NAA, which resulted in 100, 70.9 and 20.6% rooting, respectively. At least 40% of the sampled root tips had atypical starch grain deposition and abnormal graviperception. It is proposed that, at least in this clone, while IBA and NAA can be used for in vitro root induction, IAA is necessary for development of graviresponse.     

Effects of auxin and irradiance on the rooting of cuttings of Pinus sylvestris

Abstract Seedlings of Pinus sylvestris L. were grown under controlled conditions (temperature 20°C, photoperiod 17 h) at two irradiances, 8 or 40 W m^-2. Hypocotyl cuttings were excised and rooted at different irradiances in tap water solutions of indolebutyric acid (IBA). The fastest rooting and highest rooting percentage were obtained with cuttings from stock plants grown at 8 W m^-2 and treated with 10^-5M IBA for 21 days. The concentration of 10^-4M IBA inhibited root formation. In comparable treatments rooting was always better in cuttings from stock plants grown at 8 W m^-2 than in cuttings from stock plants grown at 40 W m^-2. The irradiance during the rooting period had only a minor influence on rooting. When cuttings from plants irradiated with 40 W m^-2 were treated with 10^-5M IBA for 21 days the rooting percentage almost reached the same level as in untreated cuttings from stock plants given 8 W m^-2. In cuttings treated with IBA during the whole rooting period, rooting was depressed in comparison to untreated cuttings. Aeration of the 10^-4M IBA solution increased the rooting percentage, but aeration had no effect on untreated cuttings and on cuttings treated with lower IBA concentrations.     

Shoot RNA, cambrial activity and indolebutyric acid effectivity in seasonal rooting of juvenile and mature Ficus pumila cuttings

The seasonal influence on adventitious root formation was studied in woody leaf bud cuttings of Ficus pumila L., creeping fig. Juvenile cuttings rooted easily, whereas only mature cuttings treated with indolebutyric acid (IBA) exceeded 30% rooting. Greater rooting occurred in IBA-treated juvenile and mature cuttings than controls, regardless of the month each experiment was initiated. Seasonal changes influenced rooting in all treatments except IBA treated juvenile cuttings where percentage rooting was not affected. Higher vascular cambial activity and shoot RNA levels occurred in juvenile and mature forms during peak rooting periods. Highest RNA was recorded with juvenile materials during maximum rooting periods, while lowest RNA was observed in mature shoots during low rooting intervals.