A Liquid Culture Method for the Biochemical Analysis of the Circadian Clock of Neurospora crassa

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The circadian clock that regulates the conidiation rhythm of Neurospora crassa has been reported to function normally in liquid cultures, even if they make almost no conidia and grow poorly. The phase of the rhythm was not affected by a transfer from liquid to solid medium [Perlman et al. (1981) Plant Physiol. in press]. These studies used a pantothenate-requiring auxotroph. This report describes a similar liquid culture method, in which there is no growth or conidiation and no phase shift caused by the transfer from a liquid to solid medium, and in which the wild type (bd) strain is used. Conidia were germinated in liquid medium containing glucose and arginine at the usual concentrations in continuous light. After 33 hr, discs were cut from the hyphal mats with a cork borer and transferred to liquid medium containing glucose and arginine at concentrations ten times lower than usual, then the discs were immediately placed in continuous darkness with shaking. About 18 hr after the light-dark transition, growth stopped completely and respiratory activity was suppressed due to the depletion of exogenous carbon source. No conidia were visible. But, the clock functioned normally for at least 60 hr because the phase of the rhythm of the race tubes inoculated with experimental discs was very similar to the phase of the discs which had been transferred to solid medium without culture in the low-carbon-source liquid medium. Sensitivity to perturbation by light and to cycloheximide pulse treatments also changed rhythmically. Both are evidence of normal functioning of the clock in the liquid medium. This liquid culture method will be useful for studying the biochemical mechanism of the circadian clock.
日本植物生理学会の論文

A Liquid Culture Method for the Biochemical Analysis of the Circadian Clock of Neurospora crassa

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