Abstract:

Objective: To obtain a high-yield transglutaminase (TGase)-producing strain through protoplast fusion between
mutant strains of Streptomyces mobaraensis. Methods: Protoplasts were obtained by lysozyme digestion, and after protoplast
fusion, the selected fusants were screened for TGase production by primary high-throughput screening in 96-well plates and
fermentation in test tubes and shake flasks, respectively. Results: The colonies regenerated from protoplasts had a smaller
diameter, and the diameter of colonies formed by mycelia was larger. The two different colonies had a significant difference
in TGase activity during fermentation in 96-well microplates. Different mutant strains of Streptomyces mobaraensis were
very different with respect to TGase activity, protoplast purity and protoplast regeneration rate. The maximum protoplast
regeneration rate was 1 804.25%, and the minimum was only 12.76%. After genome shuffling, we selected the top 3% of
fusant strains with high TGase activity through high-throughput screening in 96-well plates to carry out fermentation in test
tubes. Of these strains, 32.2% were high-yield strains having a 22.4% higher TGase activity than the parental strain. The
production was increased by 16.28% in shake flask fermentation. Conclusion: Genome shuffling can be used to improve
Streptomyces mobaraensis for enhanced TGase production.

Key words: Streptomyces mobaraensis, protoplasts, regeneration, fusion, genome shuffling