Rice Genetics IV - IRRI books - International Rice Research Institute

High-level expression of maize C 4-specific genes in riceRecombinant DNA techniques were used to increase the activities of C 4 photosynthesisenzymes in C 3 plants (Hudspeth et al 1992, Kogami et al 1994, Gehlen et al 1996).However, the activities of the C 4 enzymes in transgenic C 3 plants were low. In thesestudies, cDNA clones encoding the C 4 enzymes and 35S promoter from CAMV orCab promoters of target C 3 plants were used. We have demonstrated that the promotersin the maize PEPC and PPDK genes can drive the high-level expression of areporter gene in rice in a manner similar to that in maize (Matsuoka et al 1994). Thefinding that rice possesses the necessary regulatory factors for high-level expressionof the C 4 -specific genes led us to examine the possibility that intact maize C 4 -specificgenes, containing all exons and introns and their own promoter and terminator sequences,can produce high amounts of C 4 enzymes in mesophyll cells of C 3 plants.We transformed rice by introducing the intact gene for the C 4 PEPC enzyme ofmaize. More than 100 primary transformants were produced. The transgenic plantsshowed a wide range of PEPC activity. The majority (85%) had activities 2- to 30-fold higher than nontransformed plants, with the remaining 15% showing 30- to 110-fold higher activity, or 1- to 3-fold the maize PEPC activity on a protein basis (Fig. 3).Electrophoretic analysis of total leaf protein extracts revealed the presence of a novel110-kDa polypeptide, corresponding to that of maize PEPC, in transgenic rice plants.Furthermore, the amount of the polypeptide, as judged from band intensity, was wellcorrelated with the enzyme activity. Immunoblotting analysis with the antimaize PEPCantibody confirmed that the novel 110-kDa polypeptide was the product of the maizeC 4 -type PEPC gene. These results indicate that the elevated PEPC activities intransgenic rice plants were due to the expression of the maize enzyme. To our knowledge,such high-level expression of a transgene in plants has not been reported previ-Increase in PEPC activity (fold)0–1010–2020–3030–40PEPC activity in maize40–6060–8080–1100510 15 20Number of transgenic plants2530Fig. 3. The activities of PEPC in transgenic rice plants. (Note thatPEPC activity of maize genes in transgenic rice is about 35-fold higherthan that of nontransformed rice.) (From Matsuoka et al 2000.)442 Matsuoka et al