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Transgenic Res (2009) 18:1730 DOI 10.1007/s11248-008-9193-4
ORIGINAL PAPER
Genetic transformation of the sugar beet plastome
Francesca De Marchis Yongxin Wang Piergiorgio Stevanato Sergio Arcioni Michele Bellucci
Received: 3 April 2008 / Accepted: 21 May 2008 / Published online: 13 June 2008 Springer Science+Business Media B.V. 2008
Abstract It is very important for the application of chloroplast engineering to extend the range of species in which this technology can be achieved. Here, we describe the development of a chloroplast transformation system for the sugar beet (Beta vulgaris L. ssp. vulgaris, Sugar Beet Group) by biolistic bombardment of leaf petioles. Homoplasmic plastid-transformed plants of breeding line Z025 were obtained. Transformation was achieved using a vector that targets genes to the rrn16/rps12 intergenic region of the sugar beet plastome, employing the aadA gene as a selectable marker against spectinomycin and the gfp gene for visual screening of plastid transformants. gfp gene transcription and protein expression were shown in transplastomic plants. Detection of GFP in Comassie blue-stained gels suggested high GFP levels. Microscopy revealed GFP
uorescence within the chloroplasts. Our results demonstrate the feasibility of engineering the sugar beet chloroplast genome; this technology provides new opportunities for the genetic improvement of this crop and for social acceptance of genetically modied sugar beet plants.
Keywords Aminoglicoside 30-adenylyltransferase (aadA) Biolistic apparatus Chloroplast DNA
transformation Green uorescent protein
Spectinomycin Shoot regeneration
Introduction
In the past 20 years, plastids have become attractive targets in the eld of plant biotechnology. They offer considerable advantages as compared to conventional transgenic technologies, including high protein expression levels (De Cosa et al. 2001), integration into the plastome via homologous recombination without position effects or gene silencing (Daniell et al. 2001), and the expression of several transgenes in a single transcriptional unit due to the chloroplasts prokaryotic origin (Bock 2001). Moreover, from a biosafety perspective, this technology drastically reduces transgene dissemination by pollen since chloroplasts are maternally inherited in most crops (Bock and Khan 2004). Because of these advantages,
F. De Marchis Y. Wang S. Arcioni M. Bellucci (&)
Institute of Plant Genetics Research Division of Perugia, Italian National Research Council (CNR), via della Madonna Alta, 130, 06128 Perugia, Italye-mail: [email protected]
Y. WangInstitute of Animal Science, CAAS, Yuan Ming Yuan, West Road, Haidian, Beijing 100094, China
P. StevanatoDip....