Histology of the regeneration of Paulownia tomentosa (Paulowniaceae) by organogenesis / Histología de la regeneración por organogénesis en Paulownia tomentosa (Paulowniaceae)

Paulownia tomentosa is a fast-growing tree species with a considerable economic potential because of its value for wood as well as its high biomass production, and elevated stress tolerance. The objective of the present study was to evaluate the development of adventitious buds in leaves obtained from four-week-old shoots of P. tomentosa, in order to identify the cells involved in in vitro adventitious bud development. Leaves (proximal halves with the petiole) from the first node were excised from four-week-old micropropagated shoots, and cultured on Murashige and Skoog medium, supplemented with 3% (w/v) sucrose, 0.6% (w/v) Sigma agar, 22.7µM thidiazuron (TDZ) and 2.9µM indole-3-acetic acid for two weeks, explants were then transferred to the same medium with 0.44µM N6-benzyladenine for another four weeks. Five explants were collected daily during the two first weeks in TDZ treatment. A total of 140 samples were processed. Most of the buds developed indirectly from the callus formed in the petiole stub, and they became visible after eight-ten days of culture, although some buds were also observed in the area of the laminar cut at the level of the veins. The first histological changes could be observed after two-three days of culture, with the dedifferentiation of some subepidermal and inner parenchyma cells, which exhibited a large, prominent nucleus, densely-stained cytoplasm and a high nucleusto-cell area ratio. Proliferation of these cells gives rise to meristemoid formation after seven-ten days of culture. Organized cell division in meristemoids allows the formation of bud primordia that emerged from the explants surface. The progressive structural differentiation of the apical meristem, leaf primordia, and procambium strands, led to formation of complete buds that were observed in the exterior of the explants after 10-15 days of culture. Direct development of buds from cells in the subepidermic and/or epidermic layers were observed ...

Paulownia tomentosa es un árbol de rápido crecimiento y con un gran potencial económico por su madera, su utilización para la producción de biocombustible, así como su alto rendimiento en la producción de biomasa y su elevada tolerancia al estrés. El objetivo del presente trabajo ha sido evaluar el desarrollo a nivel histológico de yemas adventicias en hojas de Paulownia tomentosa. Hojas del primer entrenudo de brotes de cuatro semanas cultivados in vitro, fueron cultivadas en medio de Murashige y Skoog complementado con 22.7µM tidiazuron y 2.9µM ácido indol acético durante dos semanas. Los explantos fueron posteriormente transferidos a igual medio con 0.44µM N6 -benciladenina durante otras cuatro semanas. Se recogieron cinco muestras diarias durante las dos primeras semanas de tratamiento en medio con TDZ, procesando un total de 140 muestras. La mayoría de las yemas se desarrollan indirectamente a partir del callo formado en la superficie de corte del pecíolo. Después de dos-tres días de cultivo se observan los primeros cambios histológicos, con la desdiferenciación de algunas células de las capas subepidérmicas y del parénquima interno. La posterior proliferación de estas células da lugar a la formación de los meristemoides después de siete-diez días de cultivo. La progresiva diferenciación de estos meristemoides da lugar a la formación de las yemas que son visibles al exterior a partir de los 10-15 días. En la superficie adaxial del pecíolo se observó la formación de yemas adventicias de forma directa. Este protocolo puede ser de gran utilidad para la determinación de las células más adecuadas para los procesos de transformación genética.

In Vitro Regeneration from Shoot Tip and Nodal Explants of Simarouba Glauca DC, a Promising Biodiesel Tree.

An efficient regeneration protocol was developed from shoot tip and nodal explants of Simarouba glauca DC, a promising biodiesel plant. Nodal explants appeared to have better regeneration capacity than shoot tip explants (40%) in the tested media. The highest regeneration frequency (90%) and shoot number (7.00 ± 1.00 shoots per explants) were obtained in nodal explants in Murashige and Skoog’s (MS) medium supplemented with 6-benzylaminopurine (BAP) 4.43 μM and α-naphthalene acetic acid (NAA) 5.36 μM.Induced shoot buds were multiplied and elongated on the MS medium supplemented with BAP (4.44 μM), NAA (5.36 μM) and TDZ (Thidiazuron) 2.27 μM with 9.66±0.33 (mean length 5.35±0.32 cm) and 9.00±0.57 (mean length 4.51±0.15cm) shoots using nodal segments and shoot tip explants, respectively. Halfstrength woody plant medium (WPM) containing 2.46μM indole-3-butyric acid (IBA) produced the maximum number of roots (6.00±1.15). The rooted plantlets were hardened on MS basal liquid medium and subsequently in polycups containing sterile soil and vermiculite (1:1) and successfully established in pots.

Micropropagation and shoot regeneration of Cistus creticus ssp. Creticus.

Cistus creticus L. ssp. creticus is a medicinal aromatic shrub native in Crete (Greece). The protocol described in this paper provides optimal levels of growth regulators required to obtain high regeneration rates of Cistus in vitro. Micropropagation has been achieved through rapid proliferation of shoot-tips on Murashige and Skoog (MS) basal medium supplemented with 0.2 mg l-1 6-benzylaminopurine (BAP). After four weeks shoots were transferred to MS medium without growth regulators for further development and rooting. The highest percentage of regenerated shoots was obtained with 0.1 mg l-1 TDZ and 0.1 mg l-1 NAA after 4 weeks. Elongation and rooting was readily achieved when multiple shoots more than 1 cm in length were singled out and cultured on the MS medium without growth regulators. The plantlets were successfully adapted and grew vigorously in greenhouse conditions. This is the first report of shoot regeneration in the genus Cistus. The regeneration protocol developed in this study provides a basis for further investigation of the medicinally active constituents of this elite medicinal plant.