Culture Conditions For Plant Regeneration Research Articles

REGENERACIÓN DE PLANTAS DE Paulownia elongata Steud. POR ORGANOGÉNESIS DIRECTA


 
 
 Paulownia elongata es una especie forestal de origen oriental y rápido crecimiento, que fue introducida en México a finales de 1998 para ser incorporada en plantaciones comerciales maderables, pues su madera es ligera, por lo cual se emplea en la manufactura de muebles, artesanías, instrumentos musicales y en el acabado de interiores. El objetivo de este trabajo fue determinar las mejores condiciones de cultivo in vitro, para la regeneración de plántulas de P. elongata a partir de diversos tipos de explantes de segmentos internodales, foliares y peciolares, lo que derivó en un protocolo para organogénesis directa apto para usarse en sistemas de transformación genética. En todos los casos el medio Murashige y Skoog (MS) fue adicionado con 6-benciladenina (BA) y ácido naftalenacético (ANA), en distintas concentraciones. Se obtuvo respuesta en la totalidad de los explantes con la combinación de 5 mg L-1 de BA y 1.0 mg L-1 de ANA; sin embargo, los segmentos internodales mostraron una mejor inducción de brotes: 83% de explantes con brotes y 1.52 brotes por explante con la combinación de 4 mg L-1 de BA y 0.2 mg L-1 de ANA, a diferencia de los segmentos de pecíolo que produjeron sus mejores resultados de 53% de explantes con brote y 0.62 brotes por explante, con una mezcla de 7 mg L-1 de BA y 0.2 mg L-1 de ANA.
 
 

Optimization of culture conditions for plant regeneration ofPanicumspp. through somatic embryogenesis

AbstractWe developed a rapid and efficient shoot regeneration system forPanicumspp. by adjusting the regeneration medium and studying the responses of different genotypes and the influence of explant types (mature seed, immature embryo and shoot apex). We usedPanicum meyerianum(Nees) andPanicum longijubatum(Stapf) which were shown to perform well, to select the optimal medium for shoot regeneration. The highest frequency of shoot regeneration was obtained on Murashige and Skoog medium supplemented with 30 g L−1maltose and 1 mg L−1N‐phenyl‐N′‐[(1,2,3‐thidiazol‐5‐yl) urea]. The callus formed green spots after 1 week of culture and showed primary green shoots after 2 weeks. In this system, the calli derived from mature seed of ninePanicumgenotypes showed large variation in shoot regeneration ability: from 0 to 69.9% in the frequency of shoot formation and from 0 to 8.4 in the number of shoots per callus. Guineagrass (Panicum maximumJacq.) showed no ability and switchgrass (Panicum virgatumL.) showed low ability to regenerate from mature seed‐derived calli; however, both were able to be regenerated from immature embryos and calli derived from shoot apices. We developed an efficient protocol for high shoot regeneration of variousPanicumgenotypes which provides a foundation for efficient tissue culture and genetic improvement ofPanicum.

Improved plant regeneration and in vitro somatic embryogenesis of St Augustinegrass [Stenotaphrum secundatum (Walt.) Kuntze

AbstractSt Augustinegrass [Stenotaphrum secundatum (Walt.) Kuntze] is an important warm season turf and pasture grass. In vitro tissue culture of St Augustinegrass could serve as an important mean for its improvement through genetic transformation as well as induced somaclonal variation. To optimize tissue culture conditions for plant regeneration of St Augustinegrass, tissue culture responses of 11 explant tissues and four callus induction/subculture media have been examined. Embryogenic calli with regeneration potential were observed on cultures of early immature embryo [3 days after pollination (DAP)], immature embryo (7–14 DAP), and shoot base of young seedlings. The addition of benzyladenine (BA) in the callus induction/subculture medium enhances callus regeneration ability and does not harm callus induction for immature embryos. The best response came from 7 to 14 DAP immature embryo on MS medium containing 1 mg/l 2,4‐dichlorophenoxyacetic acid and 0.5 mg/l BA. The callus induction and regeneration rates were 97.7% and 47.6% respectively. However, BA supplement reduced callus formation and failed to enhance regeneration for young leaf bases. Scanning electron microscopy revealed that plant regeneration of St Augustinegrass is via somatic embryogenesis.

Plant Regeneration from Immature Zygotic Embryo-Derived Embryogenic Calluses and Cell Suspension Cultures of Catharanthus roseus

Culture conditions for plant regeneration in immature zygotic embryo-derived embryogenic cell suspension cultures of Catharanthus roseus (Madagascar periwinkle) ‘Little Bright Eye’ are described. Immature zygotic embryos formed off-white, friable calluses at a frequency of 20% on Murashige and Skoog (MS) medium supplemented with 4.52 µM 2,4-dichlorophenoxyacetic acid (2,4-D) after 8 weeks of culture. After a second subculture using MS basal medium at 4-week intervals, off-white friable calluses formed a small quantity of yellowish, compact embryogenic calluses. Upon transfer to MS basal medium, embryogenic calluses gave rise to numerous somatic embryos. Cell suspension cultures were established with embryogenic calluses using liquid MS medium supplemented with 4.52 µM 2,4-D. Embryogenic cell clumps from cell suspension cultures developed into plantlets at a frequency of 56.7% when plated onto MS basal medium. Plantlets were transplanted to potting soil and grown to maturity in a growth chamber.

Optimizing embryogenic callus production and plant regeneration from 'Tifton 9' bahiagrass seed explants for genetic manipulation

Bahiagrass (Paspalum notatum Flugge) is a warm season forage grass widely cultivated in southeastern U.S. and South America. The cultivar Tifton 9 has several desirable characteristics such as high forage yield, more vigor at the seedling stage, etc.; but its forage quality is very low. As an initial step for future genetic manipulations to improve its forage characteristics, we have optimized in vitro culture conditions for plant regeneration. In this report, we describe an efficient method for embryogenic callus induction and plant regeneration from bahiagrass (cv. Tifton 9) seed explants, which are readily available and easy to manipulate, compared to other explant sources reported in the literature. Murashige and Skoog (MS) medium containing 30 μM dicamba and 5 μM 6-benzyladenine (BA) was optimal for callus induction and growth. Out of 9734 seeds cultured, 65.7% germinated and 21.4% produced embryogenic callus on this medium. Shoot formation was best when embryogenic calluses induced in this medium were transferred to MS medium supplemented with 5 μM BA and 1 μM gibberellic acid with 1640 plantlets formed per gram fresh weight of callus tissue. When transferred to hormone-free SH medium, shoot systems produced well-developed root systems. The resulting plantlets grew normally produced viable seeds when transferred to soil in the greenhouse. Histochemical staining for GUS activity arising from transient expression of the introduced uidA (β-glucuronidase) gene indicated that bahiagrass embryogenic callus produced by this method is suitable for gene transfer via biolistic bombardment; and it can serve as a good target tissue for future genetic manipulations to improve the forage quality of bahiagrass (cv. Tifton 9).

Improvement of plant regeneration from long-term cultured calluses of Taipei 309, a model rice variety in in vitro studies

An efficient protocol was developed for regeneration of plants from long-term cultured calluses, which originated from mature seeds of a model rice variety Taipei 309 and were maintained by subculture for at least 6 months. The calluses were precultured for 4 weeks on a medium containing 8.88 μmol 6-benzyladenine, 5.37 μmol α-naphthaleneacetic acid and various concentrations of abscisic acid, which converted the calluses to a state more responsive to the subsequent culture conditions for plant regeneration. Supplementation of 8.69 mmol proline in the preculture medium increased the growth rate of the callus masses by 50% and resulted in the regeneration of 60% more plants. A more pronounced effect was observed after raising the 6-benzyladenine concentration to 55.48 μmol in the preculture medium, which promoted the development of adventitious buds on the calluses and led to the regeneration of some 30% more plants of better quality. Results indicate that manipulation of medium supplements and growth regulators leads to efficient plant regeneration in long-term callus cultures of rice.

Somatic embryogenesis and plant regeneration in callus culture of tef, Eragrostis tef (Zucc.) Trotter

The study was carried out to establish in vitro culture conditions for plant regeneration of tef, Eragrostis tef (Zucc.) Trotter. Mature seeds of two Ethiopian varieties, DZ-01-354 and DZ-01-196, were used to initiate callus cultures on Murashige and Skoog (MS) medium with different auxins. Four- and 8-week-old calli induced on a medium with 2.0 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D) were subcultured onto various media to induce somatic embryogenesis. Compact, nodulated, embryogenic callus was observed after transfer onto MS-callus proliferating (CP) medium. Embryogenic tissue appeared on soft and amorphous callus and developed into somatic embryos during a subsequent subculture to MS embryo-promoting (EP) media. Various growth regulator combinations were tested in CP and EP media to obtain a high efficiency of somatic embryo formation. The highest frequency of calli forming somatic embryos (56.1–68.3%) was observed when CP media with 2.0 or 4.0 mg/l 2,3,5-triiodobenzoic acid were employed and then cultures were transferred to EP media with 0.5 mg/l 2,4-D and 0.5 mg/l kinetin followed by 0.5 mg/l indole-3-acetic acid and 0.5 mg/l N6-benzyladenine. Plant development from somatic embryos was obtained on MS medium supplemented with 1.0 mg/l gibberellic acid. On average, 71.2% of calli displaying somatic embryos converted into plants. Regenerated plants were successfully transferred to soil. Neither chlorophyll-deficient plants nor morphological variants were found among regenerants. All regenerated plants were fertile.