Matthew A. Bailey

Special— Workshop: Alternative Markers for Plant Transformation Workshop Presentations from the 2000 World Congress on in vitro Biology

Marker genes are used to select and/or otherwise distinguish transgenic cells, tissues, organs or individuals from their wild-type counterparts. For several years, an impressive array of marker genes has been available for use in many diverse applications of plant genetics. Bowen (1993), for example, described 55 effective or potential selective agent/marker gene combinations representing 40 unique selectable, lethal or assayable genes. Few additional markers have been widely adopted since this review. For generation of transgenic plants, herbicide and antibiotic resistance genes have been the most utilized selectable markers. To generate an updated assessment of current selectable marker usage for production of transgenic plants, an informal survey of 47 posters, oral presentations and abstracts was conducted at the 2000 World Congress of In Vitro Biology (Table 1). While the survey was restricted to contributions at this meeting and was not exhaustive therein, it is likely to be a reasonable sample of routine, contemporary marker usage and of new marker research. Strikingly, 72% of surveyed studies utilized antibiotic (npt II, hpt) or herbicide (bar) resistance genes. Another 17% utilized gfb expression as a selection tool, but usually this marker was used in combination with antibiotic or herbicide resistance genes. The remaining contributions (11%) consisted of genes which conferred the capacity for utilization of normally non-metabolizable substrates (pmi, xyla) or which promoted preferential growth and/or morphogenesis of transformed cells (lec1, ipt, rola, b, c; Table 1). Not only were herbicide and antibiotic resistance genes most common, but also they were used effectively in .20 species, representing both monocots and dicots. Given the near ubiquity of herbicide and antibiotic resistance genes as selectable markers and the breadth of genetic backgrounds in which efficacy has been demonstrated, a reasonable interpretation of these data is that essentially universal selectable marker tools are available to generate transgenic plants. On the other hand, it is more probable that these few markers are almost exclusively utilized because of availability and convenience. When difficulties arise with their use, other components which affect the selection process are usually optimized (e.g. genotype). Alternative markers deserve consideration when selection processes are developed or optimized. First, selection mechanisms such as substrate utilization or endogenous stimulation of growth or morphogenesis could increase transformation efficiencies or extend the range of transformable genotypes. Second, a battery of efficacious marker options would enable multiple retransformations for stacking transgenes. Finally, public concern about the presence of herbicide and antibiotic resistance genes in food provides incentive to seek potentially more acceptable alternatives. This workshop was designed to present diverse approaches to develop alternative selectable markers, with examples that take advantage of alternative nutrient sources, hormone±biosynthetic genes, or visual screenable markers. Janet Reed et al. described the use of phosphomannose isomerase (pmi) as a selectable marker gene. Using mannose as a positive selective agent, transformation was shown to be efficient for several crops using either microprojectile bombardment or Agrobacterium. Anna Haldrup et al. showed that xylose isomerase (xyla) was effective as a marker gene for selection of regenerable cells of potato, tobacco and tomato incubated on medium with xylose. Ebinuma et al. described the MAT (Multi-Auto-Transformation) vector system. This approach combines the use of genes that stimulate growth and morphogenesis for positive selection of transformed cells with an excision