Poster: Emerging Technologies

Abs # 877: Development of negative selection system by using E.coli argE gene and synthetic N-acetylated phosphinothricin

Presenter:	Won-Il, Chung , wichung@mail.kaist.ac.kr
Authors	Won-Il, Chung  (A)   Woo-yong, Lee  (A)   Jong-hyeon, Jeong  (A)   Jeongmoo, Park  (A)   Seho, Hong  (A)
Affiliations:	(A): Korea Advanced Institute of Science and Technology

To develop negative selectable marker system, we modified phosphinothricin (Pt), a herbicide, to nontoxic N-acetyl phosphinothricin (N-AcPt) by organic N-acetylation method. Pt was N-acetylated and purified by vacuum evaporation of acetic acid and water. The structure of this product was revealed to be N-AcPt by NMR(nuclear magnetic resonance) spectroscopy. The produced N-AcPt was nontoxic to wild type tobacco when applied to the leaf surface. E. coli ornithine deacetylase (argE) gene which deacetylates N-acetyl amino acids, was isolated and introduced into the tobacco plant under the control of constitutive Cauliflower mosaic virus (CaMV) 35S promoter. In the transformed tobacco lines, the non-toxic precursor N-AcPt was converted to active herbicide Pt, which was sufficient to induce necrosis on the treated leaf surface. Hence, we could establish successful green-house level negative selection system by using E. coli argE gene and synthetic N-AcPt, which selectively kills transgenic tobacco over-expressing argE gene.