The production of alcohols directly from carbon dioxide by engineered cyanobacteria is an attractive technology for a sustainable future. Enhanced tolerance to the produced alcohols would be a desirable feature of the engineered cyanobacterial strains with higher alcohol productivity. We have recently obtained the mutant strains of Synechococcus elongatus PCC 7942 with higher tolerance to isopropanol using a single-cell screening system (Arai et al., Biotechnol. Bioeng., 114, 1771-1778, 2017). Among the mutant strains, SY1043 showed the highest isopropanol tolerance. Interestingly, SY1043 also showed higher tolerance to other alcohols such as ethanol and 1-butanol, however, the mechanisms involved in enhancing this alcohol tolerance were unclear. To reveal the alcohol tolerance mechanism of SY1043, we investigated the relationship between alcohol tolerance and four mutations found in SY1043 by genome resequencing analysis. Isopropanol tolerance was enhanced by amino acid substitution (Leu285Pro) in a hypothetical protein encoded by Synpcc7942_0180 of the wild type strain TA1297. TA4135, into which this mutation was introduced, showed a same tendency of tolerance to other alcohols (ethanol and 1-butanol). (C) 2017, The Society for Biotechnology, Japan. All rights reserved.