Auxin Type Research Articles

Improving the Rooting Capacity of Stem Cuttings of Virginia Pine by Severe Stumping of Parent Trees

Abstract Two experiments were conducted to develop a protocol for rooting stem cuttings from 5-year-old Virginia pine (Pinus virginiana Mill.). The first experiment tested the effects of stumping treatment (none and ¼, ½, and ¾ original height remaining), auxin type[indole-3-butyric acid or 1-napthalene acetic acid (NAA)], and auxin concentration (0, 2, 4, 6, 8, or 12 mM) on adventitious root formation. Stumping refers to the process of cutting back the stem combined with partial crown removal. The second experiment quantified the effects of crown position on the rooting of stem cuttings from stumped and nonstumped trees. Rooting percentage increased and percentage of mortality decreased as the severity of the stumping treatment increased. Auxin concentration significantly affected every rooting trait except root angle. Overall,cuttings from trees stumped to ¼ original height and treated with 4 mM NAA rooted in higher percentages (74%) and produced more primary roots (5.5) with greater total root lengths (601 mm) than the other treatments. The original crown position, particularly height, significantly affected rooting percentage. Primary needle length significantly increased as the severity of the stumping treatment increased and was positively correlated with rooting capacity. South. J. Appl. For. 30(4):182–187.

On the genetic components of the responsiveness of Kniphofia to growth regulators and sugars★

As part of an ongoing study about the genetic determinants of shoot regeneration we tested the hypothesis that the process of shoot formation and rooting in tissue culture involves groups of genetic determinants that can be discerned from each other. We used Kniphofia lines that were part of a breeding program and were crossed to produce a F1 hybrid. We used the Kniphofia propagation protocol to study the response of the parental lines to the propagation protocol and to see if we can discern some genetic insight from the F1 cross. It was found that the response to glucose was a receive type genetic response while the response to cytokinin to auxin ratio varied and dependent on the type of auxin. But in general, the response to cytokinin to auxin ratio was dominant and additive in most cases. The response to IBA in the rooting medium was semi dominant.

Somatic embryogenesis and plant regeneration from cotyledonary explants of green gram [Vigna radiata (L.) Wilezek.]—A recalcitrant grain legume

This study reports a protocol for plant regeneration from cultured explants of green gram [Vigna radiata (L.) Wilezek] via somatic embryogenesis. Somatic embryos were induced from nature cotyledons of var., TAP-7 and Pusa Baisaki when cultured on Murashige and Skoog (MS) medium fortified with different concentrations of 2,4-dichlorophenoxyacetic acid, α-naphthaleneacetic acid (NAA) and 2,4,5-trichlorophenoxyacetic acid singly or in combination with 2.22–8.88 μM N6-benzylaminopurine (BAP) or 2.32–9.38 μM kinetin. The type and concentration of auxin and plant genotype influenced the frequency of somatic embryogenesis. NAA was the most effective auxin for somatic embryo induction. The well-developed, cotyledonary shaped embryos of var. TAP-7 germinated into plantlets at a frequency of 56.6% on MS medium supplemented with 1.88 μM abscisic acid and 6.66 μM BAP. Regenerated plants were transferred to soil and grown to maturity with 90% survival. The protocol described here offers a good potential for genetic improvement using gene transfer techniques and the production of synthetic seeds of V. radiata.

Anatomical and nutritional requirements for induction and sustained growth in vitro of Cymodocea nodosa (Ucria) Ascherson

In vitro methods of plant micro-propagation are being considered as a possible solution for the decline in seagrass communities registered worldwide. To achieve successful plant micro-propagation, culture conditions are commonly adjusted empirically within almost species-specific conditions, to comply to the following three conditions: (i) culture establishment (ii) shoot production and (iii) rooting and hardening for planting in soil. Cultures of Cymodocea nodosa were established from axenic explants of the apical meristem (approx. 0.5 cm) which regenerated new leaf or produced leaf regenerating calli (5% of cultivated explants) in media containing 10 −6 M of the cytokinin analogue TDZ. Longer ramet explants, not fully axenic, containing internode with leaf and roots were also affected by 10 −6 M cytokinins and auxin type of regulators, as they promoted leaf extension (in cm), particularly GA. None of the explants progressed further to massive shoot propagation and new plantlet production. Instead, experiments made with ramet explants which simulated potential produced plantlet revealed that there seems to be a strong interaction within leaf, rhizome and root, since the carbon fixed in the leaf was rapidly translocated to the rest of the tissue (50% in the roots in a FW basis). The explants preferred ammonium and dihydrogen inorganic phosphate as a nutrient source, efficiently assimilating the former regardless of whether such were added to the underground or surface tissue. However, underground tissue was required to maintain P status in the cultivated explants.

Effect of auxins on indirect in vitro morphogenesis and expression of gusA transgene in a lectinaceous medicinal plant, Euphorbia nivulia Buch.-Ham.

Abstract The types of auxin in Murashige and Skoog (MS) medium containing N6-benzyladenine (BA) determined indirect morphogenesis, i.e. development to bipolar somatic embryos or monopolar shoots in Euphorbia nivulia Buch.-Ham. Indirect in vitro morphogenesis depended on growth regulators, explant excision period, and light. Calli induced from explants collected in March–April were superior in the induction of indirect morphogenesis to those collected in July–August. Light enforced in vitro morphogenesis, while darkness was inhibitory. The presence of kinetin in the medium also inhibited morphogenesis. Calli developed on explants collected in March–April grown on MS medium fortified with α-naphthaleneacetic acid (NAA) and BA facilitated indirect organogenesis, while those developed on medium containing 2,4-dichlorophenoxyacetic acid (2,4-D) and BA underwent somatic embryogenesis. MS medium with 13.3 μM BA and 2.69 μM NAA was the best for induction of shoots from callus, which developed a mean of 15.7 shoot...

Effect of auxins and plant oligosaccharides on root formation and elongation growth of mung bean hypocotyls

The interaction of auxins – IAA, IBA or NAA – with galactoglucomannan oligosaccharides (GGMOs) on adventitious root formation and elongation growth of mung bean hypocotyl cuttings was studied. GGMOs induced adventitious roots in the absence of auxins; however, their effect was lower compared with IBA or NAA. On the other hand, in the presence of auxins, GGMOs inhibited adventitious root induction. Their effect depended on the concentration of oligosaccharides and the type of auxin used. The highest inhibition effect of GGMOs at a concentration of 10−8 M in the presence of IBA and NAA was observed. In the presence of IAA their inhibition was non-significant in regard to the concentration. The interaction of auxins with GGMOs resulted in the formation of adventitious roots on a shorter part of hypocotyls compared with the effect of auxins alone. However, roots were induced more extensively along the hypocotyls treated with GGMOs compared with the control. GGMOs inhibited the length of induced adventitious roots in the presence of IAA, while in combination with IBA or NAA they were ineffective. The elongation of hypocotyls induced by IAA or IBA was inhibited by GGMOs, too. However, in the presence of NAA or by endogenous growth they were without any significant effect on elongation growth. These findings suggest that GGMOs in certain concentrations might inhibit rooting and the elongation process dependant on auxin used.

Influence of auxins in direct in vitro morphogenesis of Euphorbia nivulia, a lectinaceous medicinal plant

Somatic embryo (bipolar) or shoot (monopolar) morphogenesis in mesophyll cells of Euphorbia nivulia Buch.-Ham in vitro was dependent on the type of auxin supplementing Murashige and Skoog (MS) medium containing benzyladenine. Direct in vitro morphogenesis, i.e., organogenesis, and somatic embryogenesis were significantly influenced by seasonal growth of the donor plant, explant position (proximal, mid, and distal), and light. Explants collected in march/April were superior to July/August material. Proximal explants underwent morphogenesis more readily than mid- and tip-derived explants. Incubation in the light favored morphogenesis while darkness was inhibitory. Kinetin (Kn) was also inhibitory to morphogenesis. MS medium enriched with different levels of N6-benzyladenine (BA) alone, or in combination with α-naphthaleneacetic acid (NAA) or indole-3-acetic acid (IAA), induced adventitious shoots directly. Explants collected in March/April cultured on medium with 13.3 μM BA and 2.69 μM NAA developed the highest number of shoots, a mean of 15.2 shoots per proximal explant. Developed shoots rooted the best on half-strength MS medium with 2.46 μM indole-3-butyric acid, which developed a mean of 5.2 roots per shoot. Rooted healthy shoots could be transplanted to small pots, with an 80% survival rate. Addition of 2,4-dichlorophenoxyacetic acid (2.4-D) to BA-supplemented medium was obligatory to develop somatic embryos. MS medium containing 2.26 μM 2,4-D and 4.44 μM BA induced a mean of 44.8 somatic embryos per proximal explant. The embryos passed through distinct stages of embryogenesis, namely globular, heart, torpedo, and early cotyledonary. The embryos (88%) underwent maturation on half-strength MS medium with 2.89 μM gibberellic acid (GA3), and its subsequent transfer on half-strength MS basal medium in light conditions facilitated 80% conversion of embryos to plantlets. Direct shoots or embryos were originated from the mesophyll cells. Somatic embryo development was concurrent with the independent origin of vasculature in the bulbous basal portion. The survival rate of embryo-derived plants was 90%.

Macropropagation of an Endangered Medicinal Plant, Enantia Chlorantha Oliv

The effects of auxins and leaf size on the rooting of juvenile, single-node leafy stem cuttings of Enantia chlorantha Oliv. were investigated. Percentage survival of the cuttings differed among the auxins, their levels, and cutting leaf sizes. Half-leaf cuttings had the highest mean percentage survival of 70% under both naphthalene acetic acid (NAA) and the control followed by indole-3-butric acid (IBA) (60%) and the combination of IBA/NAA with 40% survival. Among full-leaf cuttings, the control treatment and the combination of IBA/NAA had a mean percentage survival of 100%, followed by IBA with 80% and NAA with 40%. Callusing of cutting was also affected by the different auxin treatments. Among the auxins and under full-leaf cuttings, the combination of IBA/NAA had the highest mean percentage of callused cuttings of 100%, followed by the control with 90%, while NAA had 40%. Callusing among the half-leaf cuttings was highest under the control and NAA (70%), while the lowest mean value of 30% was obtained under NAA. Rooting among the cuttings was affected by the IBA treatment only. Full-leaf cuttings treated with IBA at 50 ppm and the control treatment gave the highest percentage of rooted cuttings of 10% each. Auxin type, auxin concentration, leaf size, interaction between auxin type and concentration, interaction between auxin type and leaf size, and interaction between auxin concentration and leaf size had no significant effect on the percentage rooting of cuttings. Better survival and rooting rates under the control treatment, compared to those treated with rooting hormones at higher concentrations, imply that E. chlorantha can be propagated vegetatively at reduced cost, thus ensuring that this technology can be adopted with minimum capital to yield expected results.

Role of auxin and its modulators in the adventitious rooting of Eucalyptus species differing in recalcitrance

It is well established that auxins play a central role in the determination of rooting capacity, which is essential for vegetative propagation. Recent studies with apple trees have pointed to significant effects of auxin stability, wound related phenolics and ethylene production in the control of adventitious rooting. In the present study, a comparative analysis of the adventitious rooting of microcuttings of Eucalyptus saligna (easy-to-root species) and Eucalyptus globulus (difficult-to-root species) was carried out with different types of auxins, light intensities, presence or absence of apical meristem, different concentrations of phenolic compounds and presence or absence of an ethylene action inhibitor. Parameters evaluated were the percent rooting, number of roots per rooted cutting, length of longest root and mean rooting time. Results showed that auxins of intermediate stability are more favorable to rooting (particularly for the recalcitrant species), higher light intensities in the presence of exogenous auxins promote the rooting response, the absence of meristematic apex or externally supplied phenolics are not limiting for the rooting induced by exogenous auxins, and ethylene appears to play a minor role in the development of adventitious roots in microcuttings of Eucalyptus, indicating that the rhizogenic response results from direct effect of auxins.

"In vitro" Propagation of the PR 204/84 Peach Rootstock ("Prunus persica" x "P. Amygdalus") : The Effect of Auxin Type and Concentration on Rooting

"In vitro" Propagation of the PR 204/84 Peach Rootstock ("Prunus persica" x "P. Amygdalus") : The Effect of Auxin Type and Concentration on Rooting

Growth Stage, Auxin Type, and Concentration Influence Rooting of Stem Cuttings of Fraser Fir

Seven concentrations of IBA and seven concentrations of NAA plus a nonauxin control were tested over three growth stages to determine their effectiveness in promoting adventitious root formation on stem cuttings taken from 3- and 4-year-old stock plants of Fraser fir [Abies fraseri (Pursh) Poir.]. Cuttings were prepared in March (hardwood), June (softwood), or November (semi-hardwood) 2001, treated with auxin concentrations ranging from 0 to 64 mm, and placed under mist. Rooting percentage, percent mortality, number of primary roots, total root length, root system symmetry, and root angle were recorded after 16 weeks. Growth stage and auxin concentration significantly affected every rooting trait except root angle. NAA significantly increased the number of primary roots and total root length. However, auxin type did not significantly affect rooting percentage or percent mortality. The highest rooting percentages (99%) occurred when softwood cuttings were treated with 5 mm auxin, however, semi-hardwood cuttings also rooted at high percentages (90%) and had no mortality when treated with 14 mm auxin. Regardless of auxin type, the number of primary roots and total root length varied in similar patterns across concentration, although, NAA tended to induce a greater response. To root Fraser fir stem cuttings collected from 3- and 4-year-old stock plants, it is recommended that a concentration of 5 mm NAA should be used on softwood cuttings and 14 mm IBA on semi-hardwood cuttings. Chemical names used: indole-3-butyric acid (IBA); 1-naphthaleneacetic acid (NAA).

Enraizamento in vitro de marmeleiro cv. MC como porta-enxerto para a pereira e aclimatização das microestacas enraizadas

O objetivo deste trabalho foi determinar o tipo e a concentração de auxina que promova o enraizamento in vitro de marmeleiro (Cydonia oblonga Mill.) cv. MC como porta-enxerto para a pereira (Pyrus spp) e avaliar a sobrevivência das microestacas enraizadas durante a aclimatização às condições ex vitro. Os tratamentos consistiram de três tipos de auxina (ácido indolbutírico ‘AIB’, ácido naftalenoacético ‘ANA’ e ácido 3-indolacético ‘AIA’), utilizadas em cinco diferentes concentrações (0, 5, 10, 15 e 20miM). Inicialmente as microestacas foram cultivadas, durante sete dias, em meio de cultura constituído pelos sais de MS reduzidos à metade de sua concentração original, acrescido de mio-inositol (100mgL-1), sacarose (30gL-1), ágar (6gL-1) e de auxina. Após, as microestacas foram transferidas para um novo meio de cultura sem auxina. Microestacas enraizadas oriundas de vários tratamentos, indistintamente, foram aclimatizadas às condições ex vitro. O AIB, o ANA e o AIA apresentaram o mesmo efeito sobre a percentagem de enraizamento, obtendo-se o melhor resultado com 10miM; o ANA favoreceu o maior comprimento médio das raízes; a intensidade de formação de calo aumentou com a concentração de auxina, sendo maior com o AIB e o ANA; e ao fim de 30 dias de aclimatização, 65,12% das plantas sobreviveram.

Growth Stage, Auxin Type, and Concentration Influence Rooting of Virginia Pine Stem Cuttings

Seven concentrations of indole-3-butyric acid (IBA), seven concentrations of 1-naphthaleneacetic acid (NAA), and a nonauxin control were tested over three growth stages to determine their ability to promote adventitious rooting of stem cuttings from 3- and 4-year-old stock plants of virginia pine (Pinus virginiana Mill.). Cuttings were harvested September 2000 (semi-hardwood), February 2001 (hardwood), June 2001 (softwood), and October 2001 (semi-hardwood), treated with auxin concentrations ranging from 0 to 64 mm and placed under intermittent mist in a greenhouse. Rooting percentage, percent mortality, number of primary roots, total root length, root symmetry, root angle, and root diameter were assessed following 16 weeks. Growth stage affected every rooting trait measured except root symmetry and diameter. Auxin type affected total root length and root diameter, while auxin concentration affected every rooting trait except root angle. The highest predicted rooting percentages (46%) occurred when semi-hardwood cuttings were collected in September 2000 and treated with 7 mm auxin. Cuttings collected within the same growing season (2001) exhibited the highest predicted rooting percentage (33%) when softwood cuttings were treated with 6 mm auxin. Semi-hardwood cuttings rooted in 2001 produced the greatest number of roots and root lengths. Root diameter was significantly greater when NAA rather than IBA was applied, especially at higher concentrations.

Effects of Auxins and 2,3,5-Triiodobenzoic Acid on Somatic Embryo Initiation from Norway Spruce Zygotic Embryos (Picea abies)

Embryonal-suspensor mass (ESM) initiation from zygotic embryo sections was not affected by explant orientation whatever the type of external auxin used. Thus, polar auxin transport might not be directly involved in the control of somatic embryo formation. Application of 40 µM 2,3,5-triiodobenzoic acid (TIBA) suppressed ESM initiation in hypocotyl sections. This effect of TIBA mimicked that of a supraoptimal dose of α-naphtaleneacetic acid suggesting a detrimental effect of fast internal auxin accumulation on ESM initiation.

Factors Affecting the Agrobacterium-Mediated Transient Transformation of the Wetland Monocot, Typha latifolia

An Agrobacterium-mediated transformation system, using transient transformation assays, was used to evaluate conditions influencing transformation for the wetland monocot Typha latifolia. These studies were aimed at the long-term objective of evaluating candidate genes for phytoremediation. The binary plasmid vector pCAMBIA1301/EHA105, containing the β-glucuronidase coding sequence, was used in combination with factors known to affect transformation. These included callus age at the time of cocultivation with Agrobacterium tumefaciens, type and concentration of auxin for explant growth, light or dark culture environment, the presence or absence of acetosyringone (AS), explant type, explant wounding and the number of days used for cocultivation. The number of days needed for the first detection of transient expression of the β-glucuronidase gene was also examined. Three days of Agrobacterium cocultivation of 50-day-old seedling-derived calluses, grown on 20.7 µM (5 mg l−1) picloram supplemented medium, in the dark, resulted in higher levels of transient β-glucuronidase expression than were seen in calluses cultured on 4.5 or 22.6 µM (1 or 5 mg l−1) 2,4-dichlorphenoxyacetic acid containing media. The addition of 100 µM acetosyringone significantly enhanced transient β-glucuronidase activity. Wounding of explants, by cutting into two or three pieces, 3 days before cocultivation, increased expression of β-glucuronidase only in calluses cultured under light conditions. Transient β-glucuronidase expression was observed as early as 24 h after cocultivation and increased as the days post cultivation increased. The developed transient system will be used for stable transformation of Typha species.

Influence of Media Components and pH on Somatic Embryo Induction in Three Genotypes of Soybean

The influence of media components on the initiation of somatic embryogenesis in three genotypes of soybean was investigated. The following genotypes were used: Iroquois, Macon, and Savoy. Media modifications included sucrose concentration, type and concentration of auxin at two pH levels, and pH level independently. Immature cotyledons were used as the source of explant. Cotyledons were placed on a medium containing MS salts, B5 vitamins, sucrose, and auxin. Gelrite (0.2%) was used as the solidifying agent. Sucrose concentrations of 1, 2, 3, 4.5, or 6% were used. The auxins used included 2,4-dichlorophenoxyacetic acid (2,4-D) and naphthaleneacetic acid (NAA) with each at concentrations of 45.2, 90.4, 135.7, 180.9, and 226.2 μM. The pH of each the media was adjusted to either 5.7 or 7.0 with 1 N NaOH. In an additional experiment, the effect of the two pH levels, 5.7 and 7.0, was investigated independently. Overall, the frequency of somatic embryogenesis significantly varied among the different genotypes used in this study, with Iroquois showing the highest response. Frequency of somatic embryogenesis also varied in response to the different treatments used, including sucrose and auxin. The highest initiation (91.7%) and mean number of somatic embryos per responding explant (14.9) of Iroquois was observed in a medium containing 2% sucrose. The highest initiation (97.1%) and mean number of somatic embryos per responding explant (19.5) was observed in Iroquois on 135.7 μM 2,4-D and Savoy on 135.7 μM 2,4-D, respectively, for the auxin by pH level experiment. No significant differences were observed among the two pH treatments used.

Practical Factors Controlling in vitro Adventitious Root Formation from Walnut Shoot Microcuttings

Various factors were found to influence the in vitro induction and elongation of adventitious roots from walnut shoot microcuttings. Diverse walnut genotypes (Juglans regia, J. nigra × J. regia hybrids) and selected elite J. regia clones were micropropagated throughout the establishment of in vitro shoot-tip cultures. New evidence is presented here that demonstrates the importance of the genotype and juvenility of the plant material on the in vitro rooting ability. Selection of the best adapted genotypes to multiplication and rooting, and rejuvenation of mature clones through repetitive subcultures or micrografting were examined. Adult J. regia clones were rejuvenated through subsequent subcultures and their rooting was consequently improved. The same results were not accomplished by micrografting on juvenile shoots. A differential response to auxin type and concentration was observed for Juglans regia or J. nigra × J. regia clones. A short prerooting culture in multiplication medium, lowering the sucrose concentration in the root elongation medium and increasing the atmospheric carbon dioxide during the root elongation phase affected the number of shoots forming roots as well as the quality of plantlets and roots.

The role of vegetative propagation in the domestication of Pausinystalia johimbe (K. Schum), a highly threatened medicinal species of West and Central Africa

Pausinystalia johimbe (K. Schum), a tree of the Central Africa humid lowlands, is exploited for its bark to supply both export and local medicinal plant markets. The gradual destruction of the forests in which Yohimbe grows coupled with unsustainable bark harvesting methods, has resulted in local scarcity of this tree species. The study examined the amenability of P. johimbe to vegetative propagation through the rooting of leafy stem cuttings using low-cost technology polythene propagators. Using single-node leafy cuttings, three experiments were investigated in Cameroon: (i) Three propagation media (sawdust and 50:50 mixture of sand and sawdust). Initially, rooting was best in the mixed medium, but subsequently, cuttings set in the sawdust rooted better than those in sand and the mixture of sand/sawdust. However, there was no significant ( P >0.05) treatment effect on rooting percentage or on the mean number of roots per cutting. (ii) Three types of auxin at 50 μg per cutting (IAA, indole-3-acetic acid; IBA, indole-3-butyric acid and NAA, 1-naphthalene acetic acid) dissolved in 10 μl of alcohol. The control treatment received 10 μl of alcohol only. Significant differences in rooting percentage occurred after 3–4 weeks between the auxin-treated cuttings and the control. Within the same period IBA-treated cuttings rooted better than those with NAA and IBA. (iii) Four leaf areas: 0, 50, 100 and 200 cm 2 . Leafy cuttings rooted better than leafless cuttings after 4–5 weeks, with 50 cm 2 leaf area being the best from week 6. Significantly higher cutting mortality ( P <0.05) occurred for leafless cuttings than for leafy cuttings. The present study clearly indicated that P. johimbe is amenable to vegetative propagation technique using juvenile single-node leafy cuttings.

The role of vegetative propagation in the domestication of Pausinystalia johimbe (K. Schum), a highly threatened medicinal species of West and Central Africa

Pausinystalia johimbe (K. Schum), a tree of the Central Africa humid lowlands, is exploited for its bark to supply both export and local medicinal plant markets. The gradual destruction of the forests in which Yohimbe grows coupled with unsustainable bark harvesting methods, has resulted in local scarcity of this tree species. The study examined the amenability of P. johimbe to vegetative propagation through the rooting of leafy stem cuttings using low-cost technology polythene propagators. Using single-node leafy cuttings, three experiments were investigated in Cameroon: (i) Three propagation media (sawdust and 50:50 mixture of sand and sawdust). Initially, rooting was best in the mixed medium, but subsequently, cuttings set in the sawdust rooted better than those in sand and the mixture of sand/sawdust. However, there was no significant ( P>0.05) treatment effect on rooting percentage or on the mean number of roots per cutting. (ii) Three types of auxin at 50 μg per cutting (IAA, indole-3-acetic acid; IBA, indole-3-butyric acid and NAA, 1-naphthalene acetic acid) dissolved in 10 μl of alcohol. The control treatment received 10 μl of alcohol only. Significant differences in rooting percentage occurred after 3–4 weeks between the auxin-treated cuttings and the control. Within the same period IBA-treated cuttings rooted better than those with NAA and IBA. (iii) Four leaf areas: 0, 50, 100 and 200 cm 2. Leafy cuttings rooted better than leafless cuttings after 4–5 weeks, with 50 cm 2 leaf area being the best from week 6. Significantly higher cutting mortality ( P<0.05) occurred for leafless cuttings than for leafy cuttings. The present study clearly indicated that P. johimbe is amenable to vegetative propagation technique using juvenile single-node leafy cuttings.

Pepper (capsicum annuum L.) regenerants obtained by direct somatic embryogenesis fail to develop a shoot

Three auxin-type herbicides, namely 2.4-dichlorophenoxyacetic acid (2,4-D), (4-chlorophenoxy)acetic acid 2-(dimethylamino)ethyl ester (centrophenoxine), and quinolinecarboxylic acid (quinclorac) induced direct somatic embryogenesis in seed-derived zygotic embryo explants of sweet pepper (Capsicum annuum L.) when added to Murashige and Skoog medium with 200 mM sucrose. Optimum concentrations for embryogenesis induction were 0.40–0.45 mM and 1.15–1.30 μM for 2.4-D and centrophenoxine, respectively (in the presence of 5.0 gl−1 activated charcoal), or 40 μM for quinclorac (in medium without activated charcoal). Somatic embryos emerged from the epidermal and subepidermal tissues and developed on the surface of the explant. Centrophenoxine- or 2.4-D-mediated embryogenesis was accomplished from 95% of the explants in about 3 wk and, on average, six embryos were formed per explant. Induction efficieney was lower for quinelorac. Centrophenoxine-mediated embryognesis was possible in 10 pepper cultivars, the extent of the reponse-being genotype-dependent. embryos detached from the explant and transplanted onto a growth regulator-free medium germinated; however, the recovered regenerants were without a shoot, and some of them bore a single deformed cotyledon while others had no cotyledons. Regenerants lacking a shoot were generated irrespective of the auxin type applied and across all responsive genotypes investigated. Absence of a shoot, resulting from a failure in the establishment of a normal functioning apical shoot meristem, was the principal developmental disorder that precluded regeneration of normal plants via direct somatic embryogenesis. Since stem cells of the shoot meristem become established in globular and heart-stage embryos, we deduce that the absence of a shoot in germinating embryos could orginate from deviant differentiation at these early stages of embryogeny.