Expression, Purification and Characterization of Nano Luciferase in Escherichia coli

A large group of luminescence reactions catalyzed by luciferase are called bioluminescence. Since bioluminescence has been characterized as low background, high sensitivity and wide linear dynamic range. It is applied widely in sensitive molecular biology detection. In this research, we have synthesized a novel luciferase (nano luciferase) by whole gene synthesis, to make it a novel bioluminescent detection tool. Nano luciferase was cloned into a prokaryotic expression vector and expressed in an auto-induced system. Recombinant nano luciferase was purified by Ni-NTA affinity chromatography and size exclusive chromatography. The typical yield of recombinant nano luciferase was 25.3 mg from a liter culture medium with purity above 98%. Then, we tested the activity of recombinant nano luciferase, from 1.0×10-4 mg/mL to 1.0×10-7 mg/mL, nano luciferase has good linear response, the detection limit was as low as 1.0×10-7 mg/mL (around 5 pmol). The assay based on nano luciferase is ultrasensitive, good repeatable and easy operated. It would be applied as a basic tool to develop new sensitive bioluminescent probes with promising novel applications.