Plant Cell Reports

, Volume 30, Issue 1, pp 113–124

High-level accumulation of recombinant miraculin protein in transgenic tomatoes expressing a synthetic miraculin gene with optimized codon usage terminated by the native miraculin terminator

  • Kyoko Hiwasa-Tanase
  • Mpanja Nyarubona
  • Tadayoshi Hirai
  • Kazuhisa Kato
  • Takanari Ichikawa
  • Hiroshi Ezura
Original Paper

DOI: 10.1007/s00299-010-0949-y

Cite this article as:
Hiwasa-Tanase, K., Nyarubona, M., Hirai, T. et al. Plant Cell Rep (2011) 30: 113. doi:10.1007/s00299-010-0949-y

Abstract

In our previous study, a transgenic tomato line that expressed the MIR gene under control of the cauliflower mosaic virus 35S promoter and the nopaline synthase terminator (tNOS) produced the taste-modifying protein miraculin (MIR). However, the concentration of MIR in the tomatoes was lower than that in the MIR gene’s native miracle fruit. To increase MIR production, the native MIR terminator (tMIR) was used and a synthetic gene encoding MIR protein (sMIR) was designed to optimize its codon usage for tomato. Four different combinations of these genes and terminators (MIR-tNOS, MIR-tMIR, sMIR-tNOS and sMIR-tMIR) were constructed and used for transformation. The average MIR concentrations in MIR-tNOS, MIR-tMIR, sMIR-tNOS and sMIR-tMIR fruits were 131, 197, 128 and 287 μg/g fresh weight, respectively. The MIR concentrations using tMIR were higher than those using tNOS. The highest MIR accumulation was detected in sMIR-tMIR fruits. On the other hand, the MIR concentration was largely unaffected by sMIR-tNOS. The expression levels of both MIR and sMIR mRNAs terminated by tMIR tended to be higher than those terminated by tNOS. Read-through mRNA transcripts terminated by tNOS were much longer than those terminated by tMIR. These results suggest that tMIR enhances mRNA expression and permits the multiplier effect of optimized codon usage.

Keywords

Miraculin Codon optimization Miraculin terminator Transgenic tomato Read-through 

Abbreviations

GUS

β-Glucronidase

MIR

Miraculin

sMIR

Synthesized MIR

NOS

Nopaline synthase

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Kyoko Hiwasa-Tanase
    • 1
  • Mpanja Nyarubona
    • 1
  • Tadayoshi Hirai
    • 1
  • Kazuhisa Kato
    • 1
  • Takanari Ichikawa
    • 2
  • Hiroshi Ezura
    • 1
  1. 1.Graduate School of Life and Environmental SciencesUniversity of TsukubaTsukubaJapan
  2. 2.Scientific Senior Manager of Technology Center, Team Leader of Accreditation and Evaluation Team for the New UniversityOkinawa Institute of Science and Technology Promotion CorporationOnna-sonJapan

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