Abstract
The colour of firefly bioluminescence is primarily determined by the structure of the enzyme luciferase1. To date, firefly luciferase genes have been isolated from over 30 extant species producing light ranging in colour from deep-green to orange-yellow. We have reconstructed ancestral firefly luciferase genes and characterised the enzymatic properties of the recombinant proteins in order to predict ancestral firefly light emission. Results showed that the synthetic luciferase for the last common firefly ancestor exhibited green light. All known firefly species are bioluminescent in the larval stages2, with a common shared ancestor arising approximately 100 Mya3. Combined, our findings propose within the Cretaceous forest the common ancestor of contemporary fireflies emitted green light, most likely for aposematic display from nocturnal predation.