Luciferase Reporter Assay Protocol

The light produced by fireflies has become an important tool for molecular biologists. Firefly bio-luminescence -- produced by the luciferase conversion of luciferin to oxyluciferin -- was first identified, characterized and developed into the luciferase reporter assay by researchers at the University of California, San Diego, in 1986.

Definitions

Deoxyribonucleic acid (DNA) serves as the genetic material in cells. A promoter sequence of DNA controls the expression of a gene -- a sequence of DNA that produces a specific protein. As a separate piece of DNA, a plasmid contains a gene, such as luciferase.

Methods

A scientist constructs a plasmid that contains a promoter and the luciferase gene. This plasmid is transfected -- introduced biologically, chemically or mechanically -- into an animal cell, and later that cell's broken open to release any produced luciferase. Luciferin is added to this cell mix, and the amount of bio-luminescence produced by the luciferase conversion of luciferin to oxyluciferin is measured. Kits and detailed protocols are commercially available for this assay.

Uses

The amount of bio-luminescence produced in this assay is proportional to the strength of the promoter, and therefore serves as a useful method for comparing different promoters. This becomes important in industrial settings, as optimized promoters could increase the amount of protein produced in the same amount of time.

References

• "Molecular and Cell Biology": Firefly Luciferase Gene
• Clontech: Luciferase Reporter Kit Assay Manual

Resources

• "Coupled Bioluminescent Assays: Methods, Evaluations, and Applications"; Michael J. Corey; 2008.