The cloning and expression of C. thermocellum cellulases in E. coli
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The ability to harness cellulolytic enzymes, such as those found naturally in C. thermocellum, under optimal conditions is extremely promising in producing ethanol as an affordable renewable energy source. In order to do this, cellulase genes may be cloned into the easy-‐to-‐grow bacteria, E. coli. Previously, CelA and CelB were cloned, and CelA showed some enzymatic activity. In this project, CelC was successfully cloned, and the cloning of CelS was unsuccessfully attempted. Also, it was found that by altering the method of extracting the cellulases from E. coli, the activity of the cellulases (especially CelA) greatly increased. CelA also seemed to be expressed more in the E. coli, as shown by SDS-‐PAGE. Therefore, CelA is currently the ideal candidate for an endoglucanase to be used on a larger scale. Future research should focus on cloning CelS, which is an exoglucanase, and extending the enzymatic assay to include the breakdown of more crude forms of cellulose, such as filter paper.