Germination and conversion of somatic embryos derived from mature Quercus robur trees: the effects of cold storage and thidiazuron

Abstract

Somatic embryos of Quercus robur L. belonging to lines derived from mature trees were germinated in experiments designed to determine the effects of pregermination chilling and the temporary inclusion of thidiazuron (TDZ) in the germination medium. Germination response was evaluated in terms of the percentages of embryos exhibiting root-only elongation, shoot-only elongation, or both (conversion to plantlets). Conversion rates depended on genotype and were lower than those of oak zygotic embryonic axes germinated in vitro. Two months' storage of somatic embryos at 4°C between maturation and germination treatments did not influence their conversion frequency, although root-only germination was promoted in the three embryogenic lines investigated. Addition of 0.01-5.0 lM TDZ to the germination medium for 7 or 18 days induced multiple shoot formation in the epicotyl region of germinating embryos, although the 18-day treatment was detrimental with TDZ concentrations of 0.5-5.0 lM. Exposure to 0.05-0.1 lM TDZ for 7 days increased the frequency of shoot-only elongation and decreased that of root-only elongation in embryogenic lines with both high and low conversion ability, but whereas the conversion rate of the hard-to-convert line Sainza was significantly increased, that of the readily converted line B-17 was negatively affected. As a result, with 0.1 lM TDZ the total potential plant recovery of Sainza (the conversion rate plus the percentage of embryos exhibiting shoot-only germination) was 64%, approximately the same as that of B-17. Shoots excised from shoot-only germinating embryos can be elongated and rooted by proven micropropagation procedures. An additional result was the finding that adventitious bud regeneration was induced on the cotyledons of TDZ treated embryos (69% with 0.5 lM TDZ), which may prove useful for genetic transformation. This study is the first to have shown that TDZ can increase the germination efficiency of oak somatic embryos.

High-efficiency Agrobacterium-mediated transformation in Quercus robur: selection by use of a temporary immersion system and assessment by quantitative PCR

“…Line T2H was of juvenile origin (Cuenca et al 1999) and lines B13, B17, and Sainza were initiated from Q. robur trees of between 100 and 300 years old (Toribio et al 2004;Martínez et al 2008). These embryogenic lines were maintained by secondary embryogenesis with serial subculture at 5-week intervals (Valladares et al 2006).…”

“…Somatic embryos from transformed lines were germinated following the protocol developed by Martínez et al (2008). The resulting somatic plantlets were transferred to pots containing gamma sterilized peat:perlite (3:1), for acclimatization in a phytotron for 4-8 weeks, and were then grown in the greenhouse.…”

Improved organogenic capacity of shoot cultures from mature pedunculate oak trees through somatic embryogenesis as rejuvenation technique

“…These embryogenic lines had been maintained by secondary embryogenesis with sequential subculture on embryo proliferation medium at 6-week intervals since 2004 (B-17-SE) and 2005 (Sainza-BS-SE, Sainza-C-SE and CR-0-SE) (Valladares et al 2006). Embryo maturation and germination were performed following procedures previously reported for Q. robur SE (Martínez et al 2008). Briefly, prior to germination, somatic embryos were cultured for 4 weeks on maturation medium consisting of MS (Murashige and Skoog 1962) medium with half strength macronutrients and supplemented with 6% (w/v) sorbitol.…”

Biotechnological efforts for preserving and enhancing temperate hardwood tree biodiversity, health, and productivity

“…Different genotypes were used in these studies, and the effects of abscisic acid, carbohydrate source (sucrose or maltose), or osmoticum (polyethylene glycol) on somatic embryo maturation, germination and conversion into plants were investigated. Addition of 0.01-5 μM TDZ in the germination medium for somatic embryos of Q. robur induced multiple shoot formation, and adventitious bud regeneration (69%) was induced on the cotyledons of somatic embryos treated with 0.5 μM TDZ (Martinez et al 2008). Embryogenic callus was induced on immature cotyledon explants of northern red oak using MS medium containing 0.45 μM 2, 4-dichlorophenoxyacetic acid (2,4-D) (Vengadesan and Pijut 2009b).…”