Gene expression enhancement mediated by the 5′ UTR intron of the rice rubi3 gene varied remarkably among tissues in transgenic rice plants
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Introns are important sequence elements that modulate the expression of genes. Using the GUS reporter gene driven by the promoter of the rice (Oryza sativa L.) polyubiquitin rubi3 gene, we investigated the effects of the 5′ UTR intron of the rubi3 gene and the 5′ terminal 27 bp of the rubi3 coding sequence on gene expression in stably transformed rice plants. While the intron enhanced GUS gene expression, the 27-bp fused to the GUS coding sequence further augmented GUS expression level, with both varying among different tissues. The intron elevated GUS gene expression mainly at mRNA accumulation level, but also stimulated enhancement at translational level. The enhancement on mRNA accumulation, as determined by realtime quantitative RT-PCR, varied remarkably with tissue type. The augmentation by the intron at translational level also differed by tissue type, but to a lesser extent. On the other hand, the 27-bp fusion further boosted GUS protein yield without affecting mRNA accumulation level, indicating stimulation at translation level, which was also affected by tissue type. The research revealed substantial variation in the magnitudes of intron-mediated enhancement of gene expression (IME) among tissues in rice plants and the importance of using transgenic plants for IME studies.
KeywordsIME Intron rubi3 gene promoter Transgenic rice Translation
The authors thank the Plant Analysis Group at Syngenta Biotechnology, Inc., for determining the transgene copy number using realtime PCR, Dr. S. Spiker and M. Massel for assistance on using the fluorometer and phosphor imager, Dr. R. Shi for assistance on using the ABI 7900 HT Fluorescent Detection System, as well as Drs. J. Thomas, J. Shurtleff, and C. Saravitz, and J. Edwards for assistance on using the North Carolina State University Phytotron for plant growth. Authors are grateful to Drs. G. Allen, N. Allen, R. Dewey, and J. Nicholson for helpful discussions on the project. This work was supported, in part, by grants from the Center for Turfgrass Environmental Research and Education, North Carolina State University, The Consortium for Plant Biotechnology Research, Inc., and Syngenta Biotechnology, Inc.
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