The sulfolipid 6-sulfo-alpha-D-quinovosyldiacylglycerol is associated with the thylakoid membranes of many photosynthetic organisms. Previously, genes involved in sulfolipid biosynthesis have been characterized only in the purple bacterium Rhodobacter sphaeroides. Unlike plants and cyanobacteria, photosynthesis in this bacterium is anoxygenic due to the lack of a water splitting photosystem II. To test the function of sulfolipid in an organism with oxygenic photosynthesis, we isolated and inactivated a sulfolipid gene of the cyanobacterium Synechococcus sp. PCC7942. Extensive analysis of the sulfolipid-deficient null mutant revealed subtle changes in photosynthesis related biochemistry of O2. In addition, a slight increase in the variable room temperature chlorophyll fluorescence yield was observed. Regardless of these changes, it seems unlikely that sulfolipid is an essential constituent of a functional competent water oxidase or the core antenna complex of photosystem II. However, reduced growth of the mutant under phosphate-limiting conditions supports the hypothesis that sulfolipid acts as a surrogate for anionic phospholipids under phosphate-limiting growth conditions.