The actions of, and interactions between, auxins and cytokinins and their effect on in vitro rooting of selected Eucalyptus clones.
Year: 2011
Type: dissertation
Abstract: Clonal propagation of Eucalyptus spp. and its hybrids allows for competitiveness in the \ncommercial forestry industry through the propagation and preservation of superior/elite \ngenotypes. Vegetative propagation through rooted cuttings is the industry‟s standard \nand the choice of clones selected for plantations are determined by their rooting ability. \nHowever, as many potentially valuable genotypes are recalcitrant to adventitious \nrooting, micropropagation is the only effective means of propagating them. \nMicropropagation results in high plantlet yields, achieved primarily through the \nempirical use of the key plant growth regulators (PGRs) cytokinins and auxins, for \nshoot and root production, respectively. Their selection for use in vitro is driven by their \neffects on percent rooting rather than root quality. Little is known regarding the quality \nof the roots of the plantlets ex vitro, but there is some evidence that they are different \nfrom those of seedlings and cuttings. It was therefore hypothesized that the properties of \nexogenous PGRs and their interaction with other exogenous and endogenous PGRs, \ninfluenced root development and subsequent root quality. This was tested in vitro using \na good-rooting E. grandis (TAG31) and two poor-rooting E. grandis x nitens hybrid \nclones (GN155 and NH58). In the former, the auxins supplied during the pre-rooting \nculture stages (multiplication and elongation) were sufficient for 100% rooting in an \nauxin-free rooting medium. Different combinations of PGRs in the two pre-rooting \nstages, followed by rooting without auxins, revealed a direct relationship between the \nstability of the supplied auxin and the rooting ability of TAG31. Gas chromatographymass \nspectrometry (GC-MS) analyses indicated that endogenous shoot levels of indole- \n3-acetic acid (IAA) influenced graviperception. Also, low IAA content was associated \nwith atypical starch grain accumulation or its absence from root tips (53.1 nmol IAA gˉ¹ \nDW compared with 325.7 nmol IAA g-¹ DW in gravisensing roots). The specific roles \nof the natural auxins IAA and IBA on root morphogenesis were then investigated using \n2,3,5-triiodobenzoic acid (TIBA; inhibits IAA transport), ρ-chlorophenoxyisobutyric \nacid (PCIB; inhibits auxin signal transduction), and the auxin antagonist kinetin in the \nrooting medium, following root induction. After 3 weeks, the mean root diameter was \nsignificantly reduced from 552.8μm (control) to 129.2μm (with PCIB) and 278.6μm \n(with kinetin). TIBA increased root diameter to 833.4μm, decreased Δ root length, \nincreased root vasculature and resulted in agravitropism. Hence, whereas rooting could \nbe induced by IBA, IAA was necessary for the maintenance of vascular integrity and \ngraviperception. This critical role of IAA in root development is of importance as IBA, \nowing to its higher stability, has been traditionally relied upon for root induction in the \nmajority of micropropagation protocols. \nThe potential of incorporating IAA into the media formulations of in vitro protocols for \npoor-rooters that do not respond well to IAA was then investigated, using GN155 and \nNH58. While PCIB in the rooting medium of GN155 completely inhibited rooting, the \naddition of dihydroxyacetophenone (DHAP), an inhibitor of auxin conjugation, to the \nrooting medium, did not significantly increase % rooting in the presence of 0.1 mg 1ˉ¹ \nIBA (i.e. 50% rooting with 2mM DHAP and IBA, compared with 45% with IBA alone). \nThe results suggested that the inability of some eucalypts to induce roots easily in vitro \nwas not due to a deficiency in auxin signal transduction or to auxin conjugation. \nInstead, rooting was inhibited by an accumulation of kinetin within shoots during the \npre-rooting culture stages. The endogenous levels of PGRs in shoots of GN155 and \nNH58 showed a strong relationship (R² = 0.943) between the shoot kinetin:auxin and \nshoot rootability. Substituting kinetin with the relatively less stable natural cytokinin \ntrans-zeatin in the elongation stage resulted in a significant increase in % rooting in \nboth clones, from 19% to 45% (GN155) and from 31% to 52% (NH58), with 0.1 mg 1ˉ¹ \nIAA in the rooting medium. However, omitting all cytokinins from the elongation \nmedium, resulted in over 95% and 75% rooting of shoots of GN155 and NH58, \nrespectively, with 0.1 mg 1ˉ¹ IAA. \nThese results suggest that IAA is a requirement for root development and cannot be \nsubstituted by its analogues in certain root developmental events. Hence, IAA should be \nthe preferred auxin for eucalypt micropropagation. As fundamental research, the \napproach taken in this study circumvents the empirical method used in improving \nmicropropagation protocols. The importance of the properties and the interactions \nbetween endogenous and exogenous PGRs in regulating root morphogenesis, and the \npractical implications of these findings is emphasised.
Author: Muhammad Nakhooda
Language: English
Subfield: Plant Science
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W1484516954