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. 2019 Oct 30;85(22):e01516-19. doi: 10.1128/AEM.01516-19

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Copyright © 2019 Brouwers et al.

This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license.

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FIG 5 — Integration of ScMAL13 in CDFM21L.1 restores growth on maltose and maltotriose and enables native SeMALT1 and SeAGT1 characterization in knockout strains IMK820 and IMK823. (A) Characterization of S. eubayanus IMX1765 overexpressing ScMAL13 (▼) and CDFM21L.1 (▪) on SM with glucose, maltose, or maltotriose at 20°C. The OD₆₆₀ was measured (black), and sugar (black) and ethanol (red) concentrations were determined from the supernatant by HPLC. (B) Overview of constructed knockout strains. Knockouts of SeMALT1 (IMK820) and SeAGT1 (IMK823) were made with CRISPR-Cas9. Subsequently the SeSGA1 locus was replaced by ScPGK1_p-ScMAL13-ScTEF2_t using CRISPR-Cas9 in both strains, resulting in IMX1939 and IMX1940, respectively. (C) S. eubayanus strains IMK820 (▪), IMK823 (▲), IMX1939 (▼), and IMX1940 (⧫) were characterized on SM with maltose or maltotriose at 20°C. The OD₆₆₀ was measured (black), and sugar (black) and ethanol (red) concentrations were determined from the supernatant by HPLC. It is worth mentioning that the symbols corresponding to the strains IMK820, IMK823, and IMX1940 are highly overlapping and therefore might be difficult to visualize. All data represent averages and standard deviations of biological duplicates.