open access

https://hdl.handle.net/2115/48276

Development of an expression system using the heat shock protein 70 promoter in the red macroalga, Porphyra tenera

英語

Porphyra tenera

2012-01

Springer

Journal of Applied Phycology

24(1)

79-87

Porphyra is a commercially valuable source of food and drugs, and represents an important model organism for algal research. However, genetic research on P. tenera has been limited due to lack of a heterologous gene expression system. In the present study, we isolated a native promoter, the PtHSP70 promoter, for efficient expression of foreign genes in this organism. This promoter lies approximately 1 kb upstream of the coding sequence for Heat Shock Protein 70 (HSP70) and was isolated using adapter-mediated genomic PCR. Promoter activity was evaluated using the synthetic GUS gene (PyGUS) with optimized codons for Porphyra yezoensis. Interestingly, the PtHSP70 promoter allowed equivalent expression of PyGUS in both P. tenera and P. yezoensis, whereas the GAPDH promoter from P. yezoensis was not fully functional in P. tenera. These data suggest that the PtHSP70 promoter has a more conserved regulatory mechanism than the PyGAPDH promoter between these species. We also established an efficient transient transformation system for P. tenera by evaluating various transformation parameters such as quantity of gold particles, pressure of helium and vacuum, developmental stages of leafy gametophytes, and target distance. Under the optimal conditions of transient transformation, the frequency of GUS expression was determined by histochemical staining to be 30-50 cells per bombardment. Therefore, the new transient transformation system using the PtHSP70 promoter can be used for foreign gene expression in P. tenera, which may advance the development of P. tenera as a model organism.

学術雑誌論文

Accepted Manuscript

The original publication is available at www.springerlink.com

0921-8971

1573-5176

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