Abstract
To improve expression levels of recombinant proteins in plants, a new leader sequence was designed. Several elements known to enhance gene translation and/or transcription were considered, including the CaMV 35S Inr site, a CT-rich motif often shared by highly expressed plant genes and a poly(CAA) region widespread in tobamovirus and plant leaders. The effect of the synthetic leader on gusA expression was evaluated in genetically modified tobacco plants by measuring the β-glucuronidase activity and the mRNA level. When compared to the gusA leader of pBI121, the new sequence determined a 8.6-fold and a 12.5-fold increase of enzyme concentration taking into account the whole plant population or the above-average expressors, respectively. Since most pCAMBIA vectors harbour a very short 5′-UTR, identical to a fragment of the pBI121 leader, leader replacement with the sequence herein described is strongly suggested.
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Acknowledgments
We thank Dr. Gian Luca Bianchi (ERSA, Friuli Venezia Giulia, Italy) for technical assistance during real-time PCR. We also thank Dr. Dario Marchetti, Professor and Director of Tumor Biology Laboratories at LSU-Baton Rouge (Baton Rouge, LA, U.S.A.) for critically editing our work.
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De Amicis, F., Patti, T. & Marchetti, S. Improvement of the pBI121 plant expression vector by leader replacement with a sequence combining a poly(CAA) and a CT motif. Transgenic Res 16, 731–738 (2007). https://doi.org/10.1007/s11248-006-9063-x
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DOI: https://doi.org/10.1007/s11248-006-9063-x