Masahiro Ishiura, Takao Kondo, Susan S. Golden, Carl H. Johnson, and their colleagues discovered the gene cluster in 1998 and named the gene cluster kaiABC, as "kai" means “cycle” in Japanese. They generated 19 different clock mutants that were mapped to kaiA, kaiB, and kaiC genes, and successfully cloned the gene cluster in the cyanobacteria Synechococcus elongatus. Using a bacterial luciferase reporter to monitor the expression of clock-controlled gene psbAI in Synechococcus, they investigated and reported on the rescue to normal rhythmicity of long-period clock mutant C44a (with a period of 44 hours) by kaiABC. They inserted wild-type DNA through a pNIBB7942 plasmid vector into the C44a mutant, and generated clones that restored normal period (a period of 25 hours). They were eventually able to localize the gene region causing this rescue, and observed circadian rhythmicity in upstream promotor activity of kaiA and kaiB, as well as in the expression of kaiA and kaiBC messenger RNA. They determined abolishing any of the three kai genes would cause arrhythmicity in the circadian clock and reduce kaiBC promoter activity. KaiC was later found to have both autokinase and autophosphatase activity. These findings suggested that circadian rhythm was controlled by a TTFL mechanism, which is consistent with other known biological clocks.
In 2000, S. elongatus was observed in constant dark (DD) and constant light (LL). In DD, transcription and translation halted due to the absence of light but the circadian mechanism showed no significant phase shift after transitioning to constant light. In 2005, after closer examination of the KaiABC protein interactions, the phosphorylation of KaiC proved to oscillate with daily rhythms in the absence of light. In addition to the TTFL model, the PTO model was hypothesized for the KaiABC phosphorylation cycle.
Also in 2005, Nakajima et al. lysed S. elongatus and isolated KaiABC proteins. In test tubes containing only KaiABC proteins and ATP, in vitro phosphorylation of KaiC oscillated with a near 24 hour period with a slightly smaller amplitude than in vivo oscillation, proving that the KaiABC proteins are sufficient for circadian rhythm solely in the presence of ATP. Combined with the TTFL model, KaiABC as a circadian PTO was shown to be the fundamental clock regulator in S. elongatus