Transient Expression of GUS Gene Using Rubisco Gene Promoter in the Protoplasts of Porphyra yezoensis

Abstract

The transient expression of β-glucuronidase (GUS) genes in protoplasts of Porphyra yezoensis was examined using the ribulose-bisphosphate-carboxylase/oxygenase (Rubisco) gene promoter. The DNA fragments which contained the Rubisco gene promoter sequences were ligated with the GUS gene of plant vector pBI 221. When these vectors were introduced into Protoplasts, dark blue cells could be detected after the histochemical assay for GUS activity. The GUS activity was also detected by quantitative assay with a chemiluminescent substrate. The effective expressions of the GUS gene were obtained under the electroporation conditions of 200 to 300 V field strength, 47 ms pulse length and 0.03μg/ml vector DNA concentration. The agarose culture of protoplasts was more effective than the liquid culture to obtain high expression rate of the introduced GUS gene. In addition, high expression rates of introduced genes were observed at 4 to 5 days after the electroporations. These results show some of the technical conditions for the transient expression of GUS gene in P. yezoensis.