Overexpression of maize NADP-malic enzyme (ME) cDNA gene tagging with ACV5 epitope into rice (Oryza sativa L.)
2018
Bagunu, E. | Lin, H.C. | Covshoff, S. | Montecillo, F. | Anonuevo, J.J. | Mercado, M.A. | Salazar, P. | Lim, L. | Reyes, J. | Yin, X. | Quick P.
The NADP-malic enzyme (ME) is a key enzyme of the C sub 4 photosynthesis pathway in NADP-ME type C sub 4 plants such as maize. It catalyzes the decarboxylation of malate in the bundle sheath cells to supply CO sub 2 to Rubisco. The full-length cDNA encoding the maize (Zea mays) specific C₄ NADP-malic enzyme was expressed in rice (Oryza sativa L.) under the control of the bundle sheath specific Zoysia japonica PCK (ZjPCK)promoter. The gene was tagged with AcV5 epitope so that it will be detected during the analysis of immunoblotting and immunolocalization. In this study, the T₀ transgenic rice plants were characterized using molecular and biochemical techniques. PCR analysis showed 58 positive plants for the transformed gene but only 16 plants have protein expression based on immunoblotting analysis. Varying copy number of ME gene insertion was also observed in the Southern blot analysis. In the cross-section of transgenic rice leaf, immunolocalization studies using confocal microscope showed that maize NADP-ME was not only localized in bundle sheath cells but also in the mesophyll cells of transgenic rice plants, taken together, transgenic plants expressing the NADP-ME gene will be advanced for further screening to obtain homozygous lines and will later on be used for the gene-pyramiding of C sub 4- photosynthesis related genes to make a C sub 4 Rice Prototype.
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