Agrobacterium‐mediated transformation of reed (Phragmites communisTrinius) using mature seed‐derived calli

Abstract

Reed (Phragmites communis) is a potential bioenergy plant. We report on its first Agrobacterium-mediated transformation using mature seed-derived calli. The Agrobacterium strains LBA4404, EHA105, and GV3101, each harboring the binary vector pIG121Hm, were used to optimize T-DNA delivery into the reed genome. Bacterial strain, cocultivation period and acetosyringone concentration significantly influenced the T-DNA transfer. About 48% transient expression and 3.5% stable transformation were achieved when calli were infected with strain EHA105 for 10 min under 800 mbar negative pressure and cocultivated for 3 days in 200 lM acetosyringone containing medium. Putative transformants were selected in 25 mg l À1 hygromycin B. PCR, and Southern blot analysis confirmed the presence of the transgenes and their stable integration. Independent transgenic lines contained one to three copies of the transgene. Transgene expression was validated by RT-PCR and GUS staining of stems and leaves.

Agrobacterium-mediated genetic transformation of Miscanthus sinensis

“…For the inoculation and co-cultivation of Agrobacterium cells to deliver transgenes into embryogenic calli of M. sinensis, conditions with relatively high concentration of acetosyringone (400 lM) and relatively long co-cultivation period (5 days) were shown to be effective (Supplementary Table 2), when compared with the conditions for other monocotyledons (Kim et al 2007;Alves et al 2009;Li and Qu 2011;Kim et al 2013). As an example, 200 lM acetosyringone and 3 days of co-cultivation were used in a recent report for Agrobacteriummediated transformation of reed (Phragmites communis Trinius) using mature seeds-derived embryogenic calli (Kim et al 2013).…”

“…As an example, 200 lM acetosyringone and 3 days of co-cultivation were used in a recent report for Agrobacteriummediated transformation of reed (Phragmites communis Trinius) using mature seeds-derived embryogenic calli (Kim et al 2013). For the selection of transformed calli and transgenic plants, we initially used hygromycin antibiotics for the selection of transgenic plants following the transformation with pCAMBIA1301 vector.…”

Plant Responses to Extended Photosynthetically Active Radiation (EPAR)

“…UV, influences plant morphogenesis, 35 and interestingly callus growth and morphogenesis. In the investigations into plant callus growth, light treatment has been reported based on lux [73][74][75] or PAR, [76][77][78] even though the light sources might had radiated out of PAR region, e.g. UV or IR regions, that could alter callus growth and morphogenesis.…”

Enhancement of Agrobacterium-mediated transformation efficiency of model plant using quorum sensing molecule, N-3-oxo-octanoyl-l-homoserine-lactone

“…Difference letters indicate significant difference at p \ 0.05 Plant Cell Tiss Organ Cult medium containing high concentrations of phenolic compounds. Several evidences suggested that high concentrations of acetosyringone may suppress the virulence in some strain/plant species (Godwin et al 1991;Kim et al 2013). Jourbert et al (2004 found that the transformation efficiency for tobacco leaves was decreased after addition of 250 lM acetosyringone into inoculation and co-cultivation media.…”

“…Several studies indicated that the presence of phenolic compounds, such as acetosyringone and alphahydroxy-acetosyringone, is critical for integration of T-DNA into plant genome (Kim et al 2013;Subramoni et al 2014). Phenolic compounds have been shown to favor the transformation efficiency in numerous plant species, including Podophyllum hexandrum , Phragmites communis (Kim et al 2013), Fragaria vesca (Pantazis et al 2013), Withania coagulans (Mishra et al 2012), and Triticum turgidum (He et al 2010).…”