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GrainGenes Reference Report: TPJ-19-719

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Reference	TPJ-19-719
Title	Negative selection systems for transgenic barley (Hordeum vulgare L.): comparison of bacterial codA- and cytochrome P450 gene-mediated selection
Journal	The Plant Journal
Year	1999
Pages	719-726
Author	Koprek T McElroy D Louwerse J Williams-Carrier R Lemaux PG
Abstract	Summary: Efficient negative selection systems are increasingly needed for numerous applications in plant biology. In recent years, various counter-selectable genes have been tested in six dicotyledonous species, whereas there are no data available for the use of negative selection markers in monocotyledonous species. In this study, we compared the applicability and reliability of two different conditional negative selection systems in transgenic barley. The bacterial codA gene encoding cytosine deaminase, which converts the non-toxic 5-fluorocytosine (5-FC) into the toxic 5-fluorouracil (5-FU), was used for in vitro selection of germinating seedlings. Development of codA-expressing seedlings was strongly inhibited by germinating the seeds in the presence of 5-FC. For selecting plants in the greenhouse, a bacterial cytochrome P450 mono-oxygenase gene, the product of which catalyses the dealkylation of a sulfonylurea compound, R7402, into its cytotoxic metabolite, was used. T1 plants expressing the selectable marker gene showed striking morphological differences from the nontransgenic plants. In experiments with both negative selectable markers, the presence or absence of the transgene, as predicted from the physiological appearance of the plants under selection, was confirmed by PCR analysis. We demonstrate that both marker genes provide tight negative selection; however, the use of the P450 gene is more amenable to large-scale screening under greenhouse or field conditions
Keyword	[ Hide all but 1 of 35 ] appearance bacteria barley cytochrome p 450 cytotoxicity dealkylation field gene genetic markers growth hordeum vulgare hydrolases in vitro in vitro selection inhibition its marker marker genes metabolites p450 pcr phytotoxicity plant morphology reporter gene screening seed seed germination selectable marker selectable marker gene selection sulfonylurea transgene transgenic transgenic barley transgenic plant

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