Biohydrogen production from microalgae still remains to be discussed and examined more specifically, given that it is one of the most important energy carriers possessing environmental-friendly and sustainable characteristics. Although microalgae species capable of biohydrogen production do exist, Chlamydomonas reinhardtii is considered to be one of the most promising eukaryotic H-2 producers, and can serve as a model organism for such studies. Unfortunately, even if the metabolic basis and environmental conditions for this process are well defined, the sustainability of biohydrogen production is not straightforward. At this point, genetic engineering tools must be efficacious in order to enable mutant strains to reach desired amounts of biohydrogen. In this study, different light intensities, illumination patterns and Chlamydomonas strains such as CC124 and D1 protein mutant strains (D240, D239-40, D240-41) were investigated for the production of biohydrogen. The results showed that an increase in the light intensity shortened the lag phase of hydrogen production. With some minor differences, biohydrogen production was also found to be affected by the illumination pattern. On the other hand, maximum biohydrogen production was reached with a double-deletion mutant strain of D239-40, which attained a total production of 490 +/- 10 mL L-1 hydrogen and was followed by the other double-deletion mutant D240-41 that attained a total production of 388 +/- 10 mL L-1. (C) 2014 Elsevier B.V. All rights reserved.