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Title: Expression of an anthranilate synthase from maize mutant bf-1 in maize line HiII

item Pinkerton, Terrence
item Dowd, Patrick

Submitted to: American Society of Plant Biologists Annual Meeting
Publication Type: Abstract Only
Publication Acceptance Date: 7/11/2007
Publication Date: 7/11/2007
Citation: Pinkerton, T.S., Dowd, P.F. 2007. Expression of an anthranilate synthase from maize mutant bf-1 in maize line HiII. American Society of Plant Biologists Annual Meeting. Abstract No. 1439

Interpretive Summary:

Technical Abstract: Maize mutant bf-1 was one of a series of maize mutants generated by radiation from the Bikini Atoll atomic bomb test in 1946. It is characterized by blue fluorescence in seedlings and anthers under ultraviolet illumination and by mutant plants giving off a characteristic grape-like odor due to the accumulation of anthranilate-derived compounds. The mutant is also resistant to feedback regulation of tryptophan biosynthesis by tryptophan. Sequencing of cDNA from bf-1 plants showed a point mutation in a conserved chorismate binding region of the anthranilate synthase alpha subunit which caused a leucine to proline shift at amino acid residue 531. This mutation is unique compared to other observed mutations in plant anthranilate synthases that convey resistance to feedback inhibition of tryptophan biosynthesis. Escherichia coli expression of a bacterial anthranilate synthase carrying a mutation analogous to the L531P mutation enabled the bacteria to grow on minimal media in the presence of tryptophan biosynthesis feedback inhibitor 5-methyl tryptophan. The cloned bf-1 anthranilate synthase with an added c-terminal myc epitope tag has been cloned into the plant expression vector pAHC25 and transferred into maize line HiII by particle bombardment. Western blot analysis using anti-myc IgG to determine transgene expression, observations on phenotype in regenerated plants, and initial experiments exposing transgenic callus to the tryptophan biosynthesis feedback inhibitor 6-methyl anthranilate will be presented.