The regenerative capacity of mature pecan [Carya illinoinensis (Wangenh.) K. Koch] embryonic tissues was demonstrated after pretreating mature nuts to eliminate associated endogenous contaminants. Cultured cotyledon segments were induced to form adventitious roots in a medium with 50 μm NAA. A regeneration medium with 20 μm BA and 5 μm IBA stimulated prolific axillary shoot production from the embryonic axis without causing cotyledon abscission. Cotyledon retention was essential for shoot initiation and long-term development. Eighty-five percent of the shoots emerging from embryonic axes formed at the cotyledonary nodes. Thirty percent of the microshoots rooted on an auxin-free medium after preculture in a medium with 20 μm IBA. TDZ (25 μm) stimulated callus production from the cotyledonary nodes and radicles. Adventitious buds emerged on the callus surface and internally in callus. Chemical names used: a -naphthaleneacetic acid (NAA); 6-benzylaminopurine (BA); indole-3-butyric acid (IBA); N-phenyl-N'-1,2,3-thidiazol-5-ylurea (TDZ).
Ahmed A. Obeidy and M.A.L. Smith
Abba Upadhyaya, Tim D. Davis, Daksha Sankhla, and N. Sankhla
Both kinetin and BA promoted in vitro shoot formation from hypocotyl explants of Lupinus texensis Hook. placed on Murashige and Skoog (MS) medium. With either cytokinin, shoot formation was best at ≈4.5 μm. Adventitious root formation was observed only on tissue culture-derived shoots placed in MS media containing 5.4 to 54 μM NAA. IAA and IBA, at concentrations ranging from 5 to 55 μm, failed to stimulate rooting. Even at the optimal concentration of NAA, only 14% of the shoots produced roots. Thus, although hypocotyl explants readily produced shoots, adventitious root formation on these shoots occurred with relatively low frequency. Chemical names used: 6-benzylaminopnrine (BA); indole-3-acetic acid (IAA); indole-3-butyric acid (IBA); 6-furfurylaminopurine (kinetin); 1-naphthaleneacetic acid (NAA).
R.C. Apter, E.L. McWilliams, and F.T. Davies Jr.
One-node explants and one-node stem cuttings of Asian jasmine [Trachelospermum asiaticum (Siebold & Zucc.) Nakai] were rooted, respectively, in vitro [tissue culture (TC)] or by conventional macropropagation (MACRO). The TC and MACRO stem bases were then analyzed for differences in the time-course sequence of 1) root primordia initiation and development and 2) adventitious root xylem development and root-to-shoot xylem connections. Early root primordia were observed at Day 3, and, by Day 7, root-to-shoot xylem connections were equally developed in TC and MACRO systems. Continued development and emergence of adventitious roots were observed at Days 8 to 10. At Days 13 and 18, when viewed using scanning electron microscopy, TC root hairs were morphologically thicker and one-third to one-half the length of MACRO root hairs. There was no apparent difference in root-hair density. Inferior TC root-hair length may be a factor in the acclimation of TC-generated plantlets.
W. Michael Sullivan, Zhongchun Jiang, Richard J. Hull, and Carl D. Sawyer
Intraspecific variation in nitrate absorption by turfgrasses has been studied, but differences in turfgrass root morphology, which may contribute to observed variation, have not been ascertained. This information may benefit breeding programs aimed at improving the ability of turfgrasses to absorb nitrate from low fertility soils. This study quantified root morphological traits of Kentucky bluegrass (Poa pratensis L.) cultivars and their nitrate uptake rates (NUR). Tiller-generated plants were grown in silica sand, mowed weekly, and watered daily with half-strength modified Hoagland's nutrient solution containing 1 mM nitrate. When 5 months old, plants were excavated, and roots washed to remove sand. The plants were then transferred to 120-mL black bottles. After nitrate depletion of the nutrient solution was monitored for 8 consecutive days, the underground portion of each plant was separated into three parts: 1) adventitious roots, 2) fibrous roots, and 3) rhizomes. Measurements of total root length, total surface area, and average diameter were made by a scanning and image analysis system. NURs were calculated from nitrate depletion data and expressed as micromoles per plant per hour. Correlation analyses were performed on these morphological traits and NUR by the Minitab program. NUR was significantly and positively correlated with the total biomass, length, and area of the three underground parts. This was attributable mainly to fibrous roots as indicated by significant and positive correlations between NUR and the total biomass, length, area, and average diameter of fibrous roots. NUR was also positively correlated with the total biomass, length, and area of adventitious roots but negatively correlated with total biomass, area, and average diameter of rhizomes.
Renee Timmermann and M.A.L. Smith
The structure and quality of the adventitious root system formed on a microcutting stem is crucial to the successful acclimation, survival, and ultimate performance of micropropagated plants. Despite increasing evidence that the rooting method impacts on the character of the framework root system, very little research has elucidated the consequences of standard rooting methods on plant quality and performance. Root initiation on microcuttings is handled by a wide range of strategies in commercial practice. In comparison to in vitro roots, ex vitro roots have smaller root diameters, larger vascular diameters, greater length, more branch root development, and root hairs. One important microenvironmental parameter - light - was investigated to determine its contribution to root character. Typically, in vitro root systems are exposed to light throughout the root initiation period. Parallel treatments were established of in vitro light and dark rooting and ex vitro rooting of four woody species. Regardless of light exposure, the overall diameter of adventitious roots was larger for in vitro treatments than the ex vitro treatment. Vascular development was significantly more advanced ex vitro. These results suggest that light is not a major influence contributing to the differences between ex vitro and in vitro root character.
Robert M. Jetton, John Frampton, and Fred P. Hain
This study tested the effects of cutting length and auxin (NAA) concentration on adventitious root formation in softwood stem cuttings from mature eastern hemlock, Tsuga canadensis (L.) Carr., and carolina hemlock, T. caroliniana Engelm. Overall rooting percentage (41%) and percent mortality (22%) were higher for eastern hemlock compared with carolina hemlock (10% rooting and 13% mortality). Rooting percentage of each species responded differently to varying auxin concentrations (0, 1, 2, 4, 8 mm NAA). Maximum rooting (56%) for eastern hemlock occurred at 0 mm NAA; then decreased with increasing auxin concentration. Carolina hemlock rooting percentage increased from the control to a maximum (16%) at 1 mm NAA; then decreased with increasing auxin concentration. For both species, the lowest mortality occurred at the same auxin concentration as maximum rooting. The highest rates of mortality coincided with the same concentrations as the lowest rooting percentages. At all auxin concentrations, eastern hemlock had a higher number of roots and greater total root length relative to carolina hemlock. Mortality among 6-cm stem cuttings was twice that observed for 3-cm cuttings of both species. However, 6-cm cuttings of eastern hemlock that did form adventitious roots had more roots and longer total root length compared with 3-cm cuttings. Chemical name used: 1-naphthalenacetic acid.
C.F. Scagel, K. Reddy, and J.M. Armstrong
In a commercial nursery propagation system for hick's yew (Taxu×media `Hicksii'), we assessed whether or not the addition of inoculum of the vesicular-arbuscular mycorrhizal fungus (VAMF) Glomus intraradices into the rooting substrate during cutting propagation increased rooting, and how the quantity of inoculum influenced rooting. At 15 and 22 weeks (108 and 156 d) after cuttings were treated with root hormones and stuck, root initiation was higher on cuttings stuck in the rooting substrate containing VAMF inoculum. Increasing the quantity of inoculum in the rooting substrate increased root growth during the early stages of rooting. However the highest level of inoculum tested increased adventitious root initiation without increased root growth. Our results indicate that if VAMF inoculum is used during propagation from cuttings, there are optimal levels required to alter the initiation and growth of roots. For hick's yew, 1:100 or 2:100 (by volume) rates of G. intraradices in the rooting substrate increased the number of primary roots and growth of adventitious roots on cuttings above that achieved by using rooting hormone alone.
Guillermo Rodríguez, Sergio Aguilar-Espinosa, Eugenio Perez-Molphe Balch, María del Rocío Flores-Bello, Javier Farias-Larios, and José Gerardo López-Aguirre
The present work is the first report in vitro on root induction of Agave salmiana Otto, using Agrobacterium rhizogenes. Several concentrations of bacteria and acetosyringone were used, and different inoculation sites were tested, such as leaves, shaft, and root. Incubation time in darkness was 6 days. The transformed adventitious roots appeared 25 days after inoculation. The best treatment was when the shaft was inoculated with: 1 × 108 bacteria/mL and 100 μm acetosyringone; in this treatment, induction of transformed roots was 57.5% in the inoculated sites. The activity and presence of the foreign genes in the transformed roots of A. salmianawere verified as follows: 1) histochemical staining for GUS activity was determined in 80% of the tested root; and 2) molecular analysis via PCR was made to verify the presence of nptII gene and rol B gene (both were present in 60% of the tested root). This is the first report of the arbuscular mycorrhizal colonization on wild roots and transformed roots of Agavewith Glomus intraradicesSchenck and Smith. The result of the monoxenic culture was as follows: mother spore germinated 5 days; the colonization of the transformed roots was 70%. Then we proceeded to the recovery of daughter spores, in which we obtained an average 300 daughter spores per petri dish, 6 months after inoculation.
Mark C. Starrett, Frank A. Blazich, Steven R. Shafer, and Larry F. Grand
Selected isolates of Hymenoscyphus ericae (Read) Korf and Kernan, Oidiodendron griseum Robak, O. maius Barron, and a second O. Robak species were evaluated as inocula for in vitro establishment of micropropagated plantlets of Pieris floribunda (Pursh ex Sims) Benth. and Hook. Severity of shoot necrosis on microshoots differed for each isolate of Oidiodendron. Inoculation of micropropagated plantlets with isolates of H. ericae benefited initial shoot and root development on agar-solidified Woody Plant Medium (WPM) supplemented with sucrose and covered by a layer of autoclaved 1 peat: 1 vermiculite (v/v). Inoculation of microshoots with H. ericae or isolates of Oidiodendron did not stimulate production of adventitious roots.
Xiaoline Yu and Barbara M. Reed
Adventitious shoots were regenerated from stem segments or leaf discs of hazelnut (Corylus species) in vitro shoot cultures. Five to 10% of stem segments of `Nonpareil' regenerated adventitious shoots on modified MS medium and NCGR-COR medium supplemented with 200 mg·1-1 glutamine and combination of 1 or 5 μM thidiazuron (TDZ) and 0.1 μM naphthaleneacetic acid (NAA). Callus derived from stem segments of `Nonpareil', `Tonda Gentile Romana', and `Willamette' and leaf discs of `Dundee' cultured on medium with TDZ and NAA also produced shoots (buds) after transfer to NCGR-COR medium or modified MS medium with benzyladenine (BA) and indole-3-butyric acid (IBA). Adventitious roots were produced from leaf discs and stem segments on medium with NAA alone or with high levels of IBA or NAA combined with low levels of BA. Regenerated shoots of `Nonpareil' and `Willamette' were multiplied, rooted, and acclimatized in the greenhouse. This provides a starting point for improving the plant regeneration frequency to a level useful for genetic manipulation.