Background

The unicellular green alga Chlamydomonas reinhardtii is used as a model organism in various research fields including phototaxis of microorganisms, photosynthesis, and ciliary/flagellar motility (Hegemann and Berthold, 2009). A Chlamydomonas cell perceives light at its eyespot, the photoreceptive organelle observed as an orange spot located near the cell equator. The eyespot contains the photoreceptor proteins channelrhodopsins localized in the cellular membrane and the carotenoid-rich granule layers right behind the channelrhodopsins which function as a light reflector. Because of their relative position, the eyespot undergoes highly directional photoreception, and the cell can accurately detect the direction of light illumination (Foster and Smyth, 1980; Ueki et al., 2016). Upon photoreception, two flagella change their beating balance, and the cell changes its swimming direction either toward or away from the light source.

The Chlamydomonas phototactic direction (or ‘sign’) is regulated by cellular reduction-oxidation state, which is affected by cellular metabolism such as photosynthetic and respiratory activities (Wakabayashi et al., 2011). The phototactic sign thus indirectly reflects those activities in vivo. For instance, a mutant showing fast phototactic response has been shown to have high photosynthetic activity (Kim et al., 2016). In addition, for the regulation of flagellar beating for phototactic turning of the cell, flagellar dyneins should be strictly regulated (Kamiya and Witman, 1984; Okita et al., 2005; Hegemann and Berthold, 2009). Therefore, phototaxis assay contributes to a wide variety of biological researches, such as photoreception, photosynthesis, respiration, and motor proteins.

Various methods have been developed to quantify Chlamydomonas phototaxis. Mergenhagen developed an automatic assay system for phototaxis (photoaccumulation), which detects the density of cells in the light path by a photocell (Mergenhagen, 1984). Takahashi et al. developed a computer-assisted system that automatically detects the direction of cellular movement using an infrared-sensitive video camera (Takahashi et al., 1991). Comparing to those sophisticated systems with hand-made equipment, our protocol is rather simple, and can be carried out with equipment that is commercially or freely available.