Poster: Metabolic Engineering

Abs # 1005: Genetic Improvement of Caffeine Biosynthetic Pathway in Coffee Plants

Presenter:	Ogita, Shinjiro , ogita2002phd@hotmail.com
Authors	Ogita, Shinjiro  (A)   Uefuji, Hirotaka  (A)   Sano, Hiroshi  (A)
Affiliations:	(A): Nara Institute of Science and Technology

Coffee is widely preferred in the world as one of the most popular beverages. Caffeine (1, 3, 7-trimethylxanthine) is an essential ingredient of coffee beverages and has various potencies including stimulant, cardiac and diuretic. However, in sensitive individuals, it is thought to cause functional diseases and decaffeinated coffee is highly demanded. It is normally produced by chemical extraction method or supercritical extraction method but such products are poor in quality. With the aim of decreasing caffeine content, we attempted to genetically improve the caffeine biosynthetic pathway in coffee plants. A cDNA encoding a protein possessing 7-methylxanthine methyltransferase activity (theobromine synthase) was isolated, and designated as Coffea arabica 7-methylxanthine methyltransferase 1 (CaMXMT1). Using this clone, we produced transgenic coffee plants, in which expression of CaMXMT1 is supressed by RNAi or anti-sense methods. Embryogenic calli or somatic embryos of coffee plants were co-cultured with Agrobacterium tumefaciens EHA101 harboring pIG121-Hm, containing constructs of the gene-specific sequence in the sense or anti-sense configurations of CaMXMT1. Transformed embryogenic cell lines of C. arabica and C. canephora were selected on a modified 1/2 MS medium containing hygromycin. We investigated expression pattern of CaMXMT1 by RT-PCR method and found that, in the transformed cell lines, its transcripts were obviously suppressed by RNAi methods. As a result, endogenous level of caffeine in the transformed coffee cells dramatically reduced to less than a tenth level in comparison with that in non-transformed coffee cells.