Expression of a bacterial aroA mutant, aroA-M1, encoding 5-enolpyruvylshikimate-3-phosphate synthase for the production of glyphosate-resistant tobacco plants.

Glyphosate is a non-selective broad-spectrum herbicide that inhibits 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS), a key enzyme in the aromatic amino acid biosynthetic pathway in microorganisms and plants. We have previously reported a strategy for engineering glyphosate-resistant class I EPSPS based on staggered-PCR technology. Selected mutant enzymes exhibited high K(i)[glyphosate] and low K(m)[PEP] values compared to the parental enzymes from Escherichia coli ( EcaroA) and Salmonella typhimurium ( StaroA). One mutant, aroA-M1, was further engineered with a tobacco chloroplast leader sequence, and then placed in the binary vector pCAMBIA1300 for Agrobacterium-mediated gene transfer to tobacco ( Nicotiana tabacum cv. Xanthi). Transgenic plants with increased resistance to glyphosate were generated.