1. These findings demonstrate that Cel9M expression via the pCB102 plasmid confers stable endoglucanase capacity in C. butyricum. Additionally, we observed no significant change in the 24-hour OD 600 for pCB102-cel9M or CB_WT when supplied with 0.1 g/L CMC (Fig. 3f), suggesting that the presence of CMC and Cel9M reducing sugars cannot be utilized for the growth of C. butyricum strains;Zhang;OD 600 = 1.5) exhibited significant endoglucanase activity on CMC plates (Fig. 3d), while the supernatants from PYTG media only (Fig. S3) and the control strain CB_WT did not (Fig. 3d). Notably, the supernatant from pGG2121-cel9M lost its endoglucanase activity on CMC plates after twenty subcultures of one month, whereas both pCB102-cel9M and CB_INcel9M retained stable endoglucanase activity (Fig. 3d). Furthermore, the culture supernatants of pCB102-cel9M showed enhanced endoglucanase activity compared to CB_INcel9M, confirmed by Coomassie staining analysis (Fig. 3e),2011
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