Fig. 5: Identification of the functional mutations contributing to the improved HLHT tolerance in Synechococcus cells. | Nature Communications

Fig. 5: Identification of the functional mutations contributing to the improved HLHT tolerance in Synechococcus cells.

From: Engineered hypermutation adapts cyanobacterial photosynthesis to combined high light and high temperature stress

Fig. 5

a, b Illustrations of the two strategies utilized for evaluating the effects of the putative functional mutations on HLHT tolerance improvement. In strategy I, homologous fragments containing mutations (NC2-1, NC2-2, NC2-3, NC3-1, 0884, 1799, 1977, 1831-1, 0189-1, and 2215-2) were assembled in plasmids and transformed into Synechococcus cells, which would be screened under lethal light intensity and temperature of the WT control. Survival colonies could only be obtained from the transformed manipulations with mutations of NC2-1, NC2-2, or NC2-3 (a). For strategy II, the mutations were introduced into Synechococcus chromosomes through antibiotics-resistance-based selection. Meanwhile, recombinant strains carrying only the antibiotic resistance gene at the same locations (without the respective mutations) were constructed as controls. The parallel constructed strains (with or without a specific mutation) would be cultivated and compared in HLHT conditions (42 °C and 1500 μmol photons/m2/s) to evaluate the effects of the specific mutation. The comparisons between HS193 versus HS190 (with or without the NC2-1 mutation) and HS177 versus HS218 (with or without the 1831-1 mutation) are shown as examples (b). c The synergistic effects of the 1831-1 mutation and the NC2-1 mutation (HS228) on improving cell growths of the strain only carrying NC2-1 (HS229) at the conditions of 42 °C and 2500 μmol photons/m2/s. d The synergistic effects of the 0189-1 mutation and the NC2-1 mutation (HS230) on improving cell growths of the strain only carrying NC2-1 (HS231) at the conditions of 42 °C and 2500 μmol photons/m2/s. e The synergistic effects of the 0884 mutation and the NC2-1 mutation (HS241) on improving cell growths of the strain only carrying NC2-1 (HS199) at the conditions of 42 °C and 2500 μmol photons/m2/s. f The synergistic effects of the 1799 mutation and 0336 mutation (HS247) on improving cell growths of the strain only carrying 0336 mutation (HL7942) at the conditions of 42 °C and 1500 μmol photons/m2/s. Each experiment was replicated more than twice to ensure its reliability. Data are presented as mean values ± SD (n = 3 biological replicates). Source data are provided as a Source data file.

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