Discovery of photosynthesis genes through whole-genome sequencing of acetate-requiring mutants of < i > Chlamydomonas reinhardtii < /i >

by Setsuko Wakao, Patrick M. Shih, Katharine Guan, Wendy Schackwitz, Joshua Ye, Dhruv Patel, Robert M. Shih, Rachel M. Dent, Mansi Chovatia, Aditi Sharma, Joel Martin, Chia-Lin Wei, Krishna K. Niyogi Large-scale mutant libraries have been indispensable for genetic studies, and the development of next-generation genome sequencing technologies has greatly advanced efforts to analyze mutants. In this work, we sequenced the genomes of 660Chlamydomonas reinhardtii acetate-requiring mutants, part of a larger photosynthesis mutant collection previously generated by insertional mutagenesis with a linearized plasmid. We identified 554 insertion events from 509 mutants by mapping the plasmid insertion sites through paired-end sequences, in which one end aligned to the plasmid and the other to a chromosomal location. Nearly all (96%) of the events were associated with deletions, duplications, or more complex rearrangements of genomic DNA at the sites of plasmid insertion, and together with deletions that were unassociated with a plasmid insertion, 1470 genes were identified to be affected. Functional annotations of these genes were enriched in those related to photosynthesis, signaling, and tetrapyrrole synthesis as would be expected from a library enriched for photosynthesis mutants. Systematic manual analysis of the disrupted genes for each mutant generated a list of 253 higher-confidence candidate photosynthesis genes, and we experimentally validated two genes that are essential for...
Source: PLoS Genetics - Category: Genetics & Stem Cells Authors: Source Type: research