Codon-optimized gene for mutated shrimp luciferase and method for use thereof

摘要

There has been a demand for a codon-optimized gene for the mutated catalytic domain of Oplophorus luciferase, which is capable of efficiently expressing a protein both in a cultured animal cell and Escherichia coli. There has also been a demand for a substrate coelenterazine analog showing a higher activity than that of native 19 kDa protein. The invention provides a polynucleotide comprising a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 2. According to the invention, bis-coelenterazine is used as a substrate coelenterazine analog suitable for the photoprotein encoded by the polynucleotide comprising the polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 2.

主权项

1. A method for performing a luminescence reaction, which comprises contacting a protein encoded by a polynucleotide selected from the group consisting of (a) to (d) below:
(a) a polynucleotide comprising a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 2; (b) a polynucleotide comprising a polynucleotide encoding a protein consisting of a nucleotide sequence in which 1 to 20 nucleotides are deleted, substituted, inserted and/or added in the nucleotide sequence of SEQ ID NO: 2, and having a luminescent catalyst activity by using luciferin as a substrate; (c) a polynucleotide comprising a polynucleotide encoding a protein consisting of a nucleotide sequence having at least 90% identity to the nucleotide sequence of SEQ ID NO: 2, and having a luminescent catalyst activity by using luciferin as a substrate; and, (d) a polynucleotide comprising a polynucleotide encoding a protein which hybridizes to a polynucleotide consisting of a nucleotide sequence complementary to the nucleotide sequence of SEQ ID NO: 2 under high stringency conditions, and has a luminescent catalyst activity by using luciferin as a substrate, wherein the high stringency conditions are 5×SSC, 5×Denhardt's solution, 0.5% (w/v) SDS, 50% (v/v) formamide, and 50° C., with bis-coelenterazine or h-coelenterazine.