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- Author or Editor: Veronica Y. Laux x
Fluorescent proteins (FT) have become essential, biological research tools. Many novel genes have been cloned from a variety of species and modified for effective, stable, and strong expression in transgenic organisms. Although there are many applications, FT expression has been employed most commonly at the cellular level in plants. To investigate FT expression at the whole-plant level, particularly in flowers, petunia ‘Mitchell Diploid’ [MD (Petunia ×hybrida)] was genetically transformed with seven genes encoding FTs: DsRed2, E2Crimson, TurboRFP, ZsGreen1, ZsYellow1, rpulFKz1, or aeCP597. Each gene was cloned into a pHK-DEST-OE vector harboring constitutive figwort mosaic virus 35S promoter and NOS-terminator. These plasmids were individually introduced into the genome of MD by Agrobacterium tumefaciens–mediated transformation. Shoot regeneration efficiency from the cocultured explants ranged from 8.3% to 20.3%. Various intensities of red, green, and yellow fluorescence were detected from TurboRFP, ZsGreen1, and ZsYellow1-transgenic flowers, respectively, under ultraviolet light for specific excitation and emission filters. More than 70% of plants established from the regenerated shoots were confirmed as transgenic plants. Transgenic ZsGreen1 petunia generated strong, green fluorescence in all flower organs of T0 plants including petals, stigmas, styles, anthers, and filaments. Most of the chromophores were localized to the cytoplasm but also went into the nuclei of petal cells. There was a positive linear relationship (R 2 = 0.88) between the transgene expression levels and the relative fluorescent intensities of the ZsGreen1-transgenic flowers. No fluorescence was detected from the flowers of DsRed2-, E2Crimson-, rpulFKz1-, or aeCP597-transgenic petunias even though their gene transcripts were confirmed through semiquantitative reverse transcriptase-polymerase chain reaction. T1 generation ZsGreen1 plants showed green fluorescence emission from the cotyledons, hypocotyls, and radicles, which indicated stable FT expression was heritable. Four homozygous T2 inbred lines were finally selected. Throughout this study, we demonstrated that ZsGreen1 was most suitable for generating visible fluorescence in MD flowers among the seven genes tested. Thus, ZsGreen1 may have excellent potential for better utility as a sensitive selectable marker.
Coleus (Plectranthus scutellarioides) is an attractive and popular ornamental plant with propagation mainly achieved through vegetative cuttings. For commercial purposes, it is of interest to enhance the speed of establishment while maintaining high quality. Light quality has been shown to influence adventitious root development, so these experiments examined the effect of narrow-bandwidth light treatments on root growth and overall plant quality for seven coleus cultivars with vegetative cuttings in potting soil and one cultivar with shoot tip in vitro cultures onto Murashige and Skoog (MS) agar medium. During the 28 days of the propagation period, the cuttings grown under narrow-bandwidth red light (R; 663.4 nm at peak) more than doubled in the adventitious root number compared with those under blue light (B; 445.7 nm at peak) and green light (G; 530.0 nm at peak) in five cultivars. R light also increased fresh weight of the cuttings by 55.6% more than G light. In comparison, the cuttings grown under G light yielded significantly lower root and shoot dry mass than other light treatments. R light cuttings showed more dry mass content (9.63%) than those under white light (W; 437.4 nm and 559.5 nm at peak) and G light (7.85% and 5.86%, respectively). A positive correlation (R 2 = 0.598, P < 0.001) was found between the formation of adventitious roots and gained fresh weight of cuttings. R light made the reddish color of leaves significantly stronger in most cultivars, whereas the cuttings exposed to G light became less vivid compared with other light conditions. When the shoot tips were propagated in vitro onto MS medium, R light treatment initiated the root development more rapidly than other lights, with significantly greater rooting rate (20.0% and 63.6%, respectively) at day 5 and 10. The shoot tips under R light also formed significantly more roots (12.3 per cutting) than those grown under narrow-bandwidth B light (5.8 per cutting). The shoot tips showed browning at an early stage and newly emerged leaves grew very compactly under B light. The combination of red and green light (R+G) increased more than twice as much roots and dry mass compared with W light. In addition, the R+G light led to morphological changes, including larger leaves and longer petioles and internodes than those in other light treatments. The exposure to R+G+B and B light made the shoots very compact for the 28 days of in vitro culture period and significantly increased the chlorophyll contents resulting in dark green leaves.