Identification And Expression Of Circadian Clock Genes In Marine Diatoms

Circadian clock is an endogenous adaptation mechanism that has evolved from the long-term adaptation of organisms to environmental cyclical changes,and it is widely found in the biological world.Circadian clock endows organism ability to predict time and environmental changes,enabling the organism's internal rhythm to achieve time and space coordination with the external environment,increasing the environmental adaptability and competitiveness greatly.Diatoms are the most important primary producers in the ocean and play an important role in the matter cycle,energy flow and information transmission of marine ecosystem.In the past half century,much effort has been devoted to the circadian clocks of some model species such as Cyanophyta,Chlamydomonas,Drosophila,Arabidopsis thaliana,Rats and human beings.Composition and regulation mechanism of circadian clock have been clearly understood.However,our understanding on the circadian clock of marine diatoms,an important biological group,is almost blank.Many questions need to be studied urgently.This study selected Skeletonema costatum and Phaeodactylum tricornutum as the model species,and the known clock genes as reference genes.The bioinformatics and molecular biological methods were applied to identify circadian clock genes of marine diatoms.The circadian clock model of two marine diatom species was also established.The physiological and biochemical responses of marine diatoms and the expression patterns of circadian clock genes under different environmental conditions were studied.The main results were as follows:(1)18 and 17 circadian clock genes were identified in S.costatum and P.tricornutum,respectively.Although marine diatoms are far from higher plants such as Arabidopsis in the evolutionary status,similar circadian genes were identified in marine diatoms.Most of clock genes were same except for individual genes.Three clock core genes,CCA1,LHY and TOC1 were identified in two marine diatom species and molecular model ofmarine diatom circadian clock were established.(2)Bioinformatic analysis was used to compare the sequence of circadian clockgenes in two marine diatoms with other plant circadian clock genes and then phylogenetic trees were constructed.The core genes of CCA1,LHY and TOC1 were homologous with the core genes of other high plants.We presumed that they played a key component of oscillator in marine diatoms and were involved in the regulation of circadian clock.However,the evolutionary status of three core genes was far from A.thaliana and they might have different functions compared with higher plants.(3)The qRT-PCR technique was used to investiate the expressions ofCCA1,LHY,and TOC1 in two marine diatoms under different environmental conditions.Under normal culture conditions,three core genes exhibited 24 h rhythmical expression patterns.Three core genes reached their peaks at night,but the expression patterns differed from A.thaliana.Changes in photoperiod,nutrient concentrations such as nitrogen and phosphorus shifted the expression phase of circadian clock genes in S.costatum and P.tricornutum.However,the expression patterns between the two diatoms under changing environmental conditions were different.