Photosynthetic organisms provide food and energy for nearly all life on Earth, yet half of their protein-coding genes remain uncharacterized1,2. Characterization of these genes could be greatly accelerated by new genetic resources for unicellular organisms. Here we generated a genome-wide, indexed library of mapped insertion mutants for the unicellular alga Chlamydomonas reinhardtii. The 62,389 mutants in the library, covering 83% of nuclear protein-coding genes, are available to the community. Each mutant contains unique DNA barcodes, allowing the collection to be screened as a pool. We performed a genome-wide survey of genes required for photosynthesis, which identified 303 candidate genes. Characterization of one of these genes, the conserved predicted phosphatase-encoding gene CPL3, showed that it is important for accumulation of multiple photosynthetic protein complexes. Notably, 21 of the 43 higher-confidence genes are novel, opening new opportunities for advances in understanding of this biogeochemically fundamental process. This library will accelerate the characterization of thousands of genes in algae, plants, and animals.
Bibliographical noteFunding Information:
We thank O. Vallon for helpful discussions; M. Cahn and G. Huntress for developing and improving the CLiP website; X. Ji at the Stanford Functional Genomics Facility and Z. Weng at the Stanford Center for Genomics and Personalized Medicine for deep sequencing services; A. Itakura for help in library pooling; S. Ghosh, K. Mendoza, M. LaVoie, L. Galhardo, X. Li, Y. Wang, and Q. Chen for technical assistance; K. Barton, W. Briggs, and Z.-Y. Wang for providing lab space; J. Ecker, L. Freeman Rosenzweig, and M. Kafri for constructive suggestions on the manuscript; and the Princeton Mass Spectrometry Facility for proteomics services. This project was supported by a grant from the National Science Foundation (MCB-1146621) awarded to M.C.J. and A.R.G., grants from the National Institutes of Health (DP2-GM-119137) and the Simons Foundation and Howard Hughes Medical Institute (55108535) awarded to M.C.J., a German Academic Exchange Service (DAAD) research fellowship to F.F., Simons Foundation fellowships of the Life Sciences Research Foundation to R.E.J. and J.V.-B., an EMBO long-term fellowship (ALTF 1450-2014 and ALTF 563-2013) to J.V.-B and S.R., a Swiss National Science Foundation Advanced PostDoc Mobility Fellowship (P2GEP3_148531) to S.R., and a Westlake University startup fund to X.L.