transgenic ostreococcus tauri lines4 were cultured in keller media - supplemented artificial sea water ( km ) under 12:12 h blue ( ocean blue , lee lighting filter 724 ) light : dark cycles ( 17.5 e / m ) . for imaging , cultures were transferred to 96-well microplates ( lumitrac , greiner bio - one ) at a density of ~1510 cells / ml and entrained for 7 - 10 days . no density effects on clock output were observed under relevant density ranges , and cell division in microplates was found to be close to zero . one day prior to recording , 150 l km was replaced with 150 l km containing 333 m luciferin ( km+ ) . drugs were made up in dmso or km+ , diluted in km+ and added to replicates of 8 or 16 wells immediately prior to recording . for incubations in constant darkness , km+ was supplemented with 200 mm sorbitol and 0.4% glycerol in order to increase cell viability . bioluminescent recordings were performed on a topcount ( packard ) under constant darkness or constant red+blue led light ( 5 - 12 e / m ) . for wash - off of reversible inhibitors , cell aggregates formed in the bottom of the wells were quickly and gently washed twice with km+ , using multi - channel pipettes , and returned to recording conditions . analysis of period was performed with fft - nlls ( brass 330 ) using time windows 3 days ; mfourfit ( brass 3 ) was used to assess phase and confirmed manually . statistical analysis was performed using graphpad prism . for de novo rna synthesis analysis by [ p]utp uptake , 1 ml cell aliquots were either incubated in darkness or light / dark cycles for 4 days . 0.2 mbq of [ p]utp was added , and after incubation cells were collected and washed twice with km . sequence alignments were performed using ebi jalview , blast searches were performed using ncbi blastp under the default blossum62 settings . circadian rhythms in a eukaryote can be sustained solely by non - transcriptional mechanisms , which are conserved across taxa .
circadian rhythms are ubiquitous in eukaryotes , and co - ordinate numerous aspects of behaviour , physiology and metabolism , from sleep / wake cycles in mammals to growth and photosynthesis in plants1,2 . 
 this daily timekeeping is thought to be driven by transcriptional / translational feedback loops , whereby rhythmic expression of clock gene products regulates expression of associated genes in approximately 24-hour cycles . 
 the specific transcriptional components differ between phylogenetic kingdoms3 . 
 the unicellular pico - eukaryotic alga , ostreococcus tauri , possesses a naturally minimised clock , which includes many features that are shared with higher eukaryotes ( plants ) , such as a central negative feedback loop that involves the morning - expressed cca1 and evening - expressed toc1 genes4 . 
 given that recent observations in animals and plants have revealed prominent post - translational contributions to timekeeping5 , a reappraisal of the transcriptional contribution to oscillator function is overdue . 
 here we show that non - transcriptional mechanisms are sufficient to sustain circadian timekeeping in the eukaryotic lineage , though they normally function in conjunction with transcriptional components . 
 we identify oxidation of peroxiredoxin proteins as a transcription - independent rhythmic biomarker , which is also rhythmic in mammals6 . 
 moreover we show that pharmacological modulators of the mammalian clockwork have the same effects on rhythms in ostreococcus . 
 post - translational mechanisms , and at least one rhythmic marker , appear to be better conserved than transcriptional clock regulators . 
 it is plausible that the oldest oscillator components are non - transcriptional in nature , as in cyanobacteria7 , and are conserved across kingdoms .