Identification and Characterization of Inclusion Bodies Found in pUC19 Transformed Escherichia coli DH5α

Abstract

Inclusion bodies are protein aggregates found in various types of bacterial cells as a result of overexpression, mutations that affect the folding pathway, or exposure of cells to certain environmental stresses. Previous studies have shown that inclusion bodies are formed when Escherichia coli DH5 α were transformed with plasmid pUC19. We hypothesized that the inclusion bodies observed were due to overexpression of plasmid genes, specifically lacZ’ R which encodes the amino-terminal fragment of β -galactosidase, and amp which encodes β – lactamase. To test the hypothesis, we attempted to purify these inclusion bodies and determine their composite proteins via western blotting. Based upon the results obtained, we were unable to show that any inclusion bodies were formed due to aggregation of β – galactosidase and β -lactamase. Furthermore, it is highly possible that no inclusion bodies were present at all due to the lack of distinctive protein band patterns in the inclusion bodycontaining samples. However, the purification procedures used were crude and were not optimized; hence inclusion bodies, if present, may not have been separated from other components in the cells. Thus the possibility of inclusion bodies being present still exists.