分子生物学文献.pdf

Journal of Biotechnology 121 2006 309–317 Trehalose-6-phosphate synthase as an intrinsic selection marker for plant transformation Barbara Leyman a,?, Nelson Avonceb , Matthew Ramon a , Patrick Van Dijck a , Gabriel Iturriaga b , Johan M. Thevelein a a Department of Molecular Microbiology, Flanders Interuniversity Institute for Biotechnology VIB and Laboratory of Molecular Cell Biology, Institute of Botany and Microbiology, Katholieke Universiteit Leuven, Kasteelpark Arenberg 31, B-3001 Leuven-Heverlee, Flanders, Belgium b Centro de Investigaci′on en Biotecnolog′?a-UAEM, Universidad 1001, Col. Chamilpa, Cuernavaca 62210, Mexico Received 12 April 2005; received in revised form 9 August 2005; accepted 24 August 2005 Abstract Insertion of foreign DNA into plant genomes occurs randomly and with low frequency. Hence, a selectable marker is generally required to identify transgenic plants. Until now, all selection systems have been based on the use of non-plant genes, derived from microorganisms and usually conferring antibiotic or herbicide resistance. The use of microorganism-derived genes however has raised biosafety concerns. We have developed a novel selection system based on enhancing the expression of a plant-intrinsic gene and the use of a harmless selection agent. Selection takes advantage of the reduced glucose sensitivity of seedlings with enhanced expression of AtTPS1 , a gene encoding trehalose-6-P synthase. As a result, transformants can be identi?ed as developing green seedlings amongst the background of small, pale non-transformed plantlets on high glucose medium. In addition, vegetative regeneration of tobacco leaf explants is very sensitive to high external glucose. Overexpression of AtTPS1 in tobacco allows selecting gluc