However, we must point out that
the BX-795 in vitro host strain used to generate the stm6 mutant is a low H2 producer compared to other Chlamydomonas WT strains such as CC-124 and D66. It would be more useful if the stm6 mutant genotype were genetically transferred to one of these high H2-producing WT strains to increase the chance that it will achieve higher conversion efficiencies in the future. Barrier: photosynthetic efficiency The concept of decreasing the chlorophyll antenna size of the photosystems to increase the light utilization efficiency of algal mass cultures has been proposed in the past (Melis et al. 2000; Melis and Chen 2005). Research efforts to test it have focused on using random mutagenesis and high-throughput screening to aid the identification of genes that regulate the Chl antenna size in green alga. This work has resulted in strains with gradually smaller Dinaciclib antenna sizes and increasing photosynthetic productivity (Polle et al. 2003; Tetali et al. 2007; Mitra and Melis
2010; Kirst et al. 2012a, b). Analysis of the Chlamydomonas tla1 truncated antenna mutant proved that the concept is also successful in increasing H2 productivity. Kosourov et al. 2011 immobilized WT and tla1 sulfur-deprived mutant cells on alginate fims and monitored long-term H2-photoproduction activity under light intensities ranging from 19 to 350 μE m−2 s−1PAR. They showed that the mutant was able to produce H2 gas for over 250 h under all light conditions tested and exhibited a 4–8 times higher maximum specific rate between 285 and 350 μE m−2 s−1, compared to WT cells. Along the same line, RNAi knockdowns of the light-harvesting complexes M1, 2, and 3 were performed to reduce the antenna size and optimize light capture by Chlamydomonas. LHCBM1, 2, and 3 are known to be the most abundant Metalloexopeptidase LHC proteins, and knocking them down simultaneously reduced the total chlorophyll content of the cells—resulting in improved light penetration and utilization. This multiple mutant displayed higher
photosynthesis light saturation level and did not suffer photoinhibition under saturating light intensity. Upon sulfur deprivation, the mutant strain showed an immediate onset of H2 production, indicating that the intracellular O2 levels were already poised to induce HYDA transcription. Furthermore, the rate of H2 production observed in this strain was twice as high as that of the stm6GLC4 (Oey et al. 2013) described below. As mentioned in the previous section, both the tla and the lhcb mutants are being or have been introduced into strains that are not limited by the non-dissipation of the proton gtradient and will continue to serve as the host for other strains expressing additional useful traits.