High-level expression of maize phosphoenolpyruvate carboxylase in transgenic rice plants

Abstract

Using an Agrobacterium-mediated transformation system, we have introduced the intact gene of maize phosphoenolpyruvate carboxylase (PEPC), which catalyzes the initial fixation of atmospheric CO2 in C4 plants into the C3 crop rice. Most transgenic rice plants showed high-level expression of the maize gene; the activities of PEPC in leaves of some transgenic plants were two- to threefold higher than those in maize, and the enzyme accounted for up to 12% of the total leaf soluble protein. RNA gel blot and Southern blot analyses showed that the level of expression of the maize PEPC in transgenic rice plants correlated with the amount of transcript and the copy number of the inserted maize gene. Physiologically, the transgenic plants exhibited reduced O2 inhibition of photosynthesis and photosynthetic rates comparable to those of untransformed plants. The results demonstrate a successful strategy for installing the key biochemical component of the C4 pathway of photosynthesis in C3 plants.

Transformation and characterization of transgenic rice and Cleome spinosa plants carrying the maize phosphoenolpyruvate carboxylase genomic DNA

“…No apparent differences in phenotype were observed in any of the transgenic plants as compared to their wildtype parents. This result is similar to other reports that over 90% transgenic rice plants carrying the same construct in different cultivars exhibited normal phenotype as the wild-type origin (Ku et al 1999;Bandyopadhyay et al 2007).…”

“…The enzyme activities of PPDK and PEPC in these wheat lines were only about twofold and fourfold, respectively, higher than those of the control plants. These expression levels are much lower than those achieved in the transgenic rice plants (Ku et al 1999;Taniguchi et al 2008). Whether the differences in expression levels of these maize genes in the C 3 rice and wheat affect their photosynthetic performance and other physiological traits under different conditions remain to be investigated.…”

“…Protein extraction, gel electrophoresis, western blot analysis, and assay of PEPC activity were as previously described (Ku et al 1999). Briefly, 0.1 g leaf tissue from wild-type and transgenic plants was harvested and quickly ground in 1.5 ml extraction buffer (50 mM Tris-HCl, pH 7.0; 10 mM MgCl 2 ; 1 mM EDTA; 5 mM dithiothreitol; 5% polyvinylpolypyrrolidone; 10% glycerol) using a cold pestle and mortar.…”

“…Previously, the transformation vector PEPCgenome/pBIH2 carrying the XbaI fragment (8.8 kb) of maize pepc gene (including all exons and introns, promoter and terminator sequences) and two selective antibiotic resistant genes neomycin phosphotransferase II (nptII) and hygromycin phosphotransferase II (hptII) was used to transform two japonica rice cultivars (Kitaake and Nipponbare) via an Agrobacterium-mediated method. Most of the transgenic rice plants generated showed high-level expression of the maize pepc gene and exhibited reduced sensitivity of photosynthesis to O 2 inhibition (Ku et al 1999). Later, the same construct was used to transform indica rice IR68899B using a bombardment method, and the transformants showed enhancement in photosynthesis rate under high temperature (Bandyopadhyay et al 2007).…”

Utility of basic research in plant/crop physiology in relation to crop improvement: a review and a personal account

“…Noteworthy in this regard is the research on molecular engineering of C 4 photosynthesis (Matsuoka et al, 1994(Matsuoka et al, , 2001) and the attempt to transform C 3 rice by inserting the gene controlling C 4 PEPC from maize (Ku et al, 1999). According to Ku (http:// www.biotech-info.net), such transgenic rice plants showed a high expression of PEPC within leaves, greater grain yield, and higher leaf photosynthetic rate per unit leaf area.…”