A novel principle for selection of transgenic plant cells: Positive selection

A novel principle for selection of transgenic plant cells is presented. In contrast to traditional selection where the transgenic cells acquire the ability to survive on selective media while the non-transgenic cells are killed (negative selection), this selection method actively favours regeneration and growth of the transgenic cells while the non-transgenic cells are starved but not killed. Therefore, this selection strategy is termed 'positive selection'. The E. coli beta-glucuronidase gene was used as selectable (as well as screenable) gene and a glucuronide derivative of the cytokinin benzyladenine as selective agent which is inactive as cytokinin but, upon hydrolysis by GUS, active cytokinin is released stimulating the transformed cells to regenerate. Selection of Agrobacterium tumefaciens inoculated of tobacco leaf discs on benzyladenine N-3-glucuronide (7.5-15 mg/l) resulted in 1.7-2.9 fold higher transformation frequencies compared to kanamycin selection. A significant advantage of this selection procedure is the elimination of the need for herbicide and antibiotic resistance genes.