A heterotrophic synchronous culture of Chlorella

Based on the observation that photoautotrophically grown Chlorella cells fail to complete cell division under anaerobic condition in the dark, we devised a heterotrophic synchronous culture (HSC) system for this green alga. The system consists, at a temperature of 21 degrees C, of a 32 +/- 1 h period under 3% CO2 in air and the successive 12 +/- 1 h period under 0.5% O2 +/- 99.5% N2 (the sum of the two periods is equal to 44 +/- 1 h), using a semi-balanced medium containing glucose in an inorganic salt solution. This procedure could be successively repeated several times and the division index was practically 100%, as it is for Chlorella in the photoautotrophic synchronous culture (PSC). We believe this is the first successful HSC achieved by the method of alternating atmospheric oxygen tension. Synthetic patterns for DNA, RNA and protein as well as chlorophyll during the cell cycle of Chlorella in HSC were similar to those observed in PSC, and the respiratory activity of actively growing cells in HSC was twice that of those in PSC. It is hoped that this system, under the regimen of high and low oxygen tension, can be utilized for other eukaryotic cells.