Microbial Cell Culture

Cloning and Recombinant Protein Expression

Bacterial cultures are used for recombinant protein expression, plasmid cloning and amplification, and bacterial artificial chromosome (BAC) and cosmid cloning. Bacterial cells can be altered to uptake and incorporate exogenous genes and plasmids through transformation. Transformation of competent cells typically occurs through electroporation or heat shock of chemically-treated competent cells.

Transformed bacteria are then identified and selected based on acquired antibiotic resistance imparted by recombinant plasmid DNA or using “blue-white” screening to identify recombinant bacteria expressing β-galactosidase as a marker. Yeast can also be transformed using electroporation and other methods for yeast two-hybrid and reporter gene assays to study protein expression and protein interactions.

Cultivation for Microbial Isolation and Identification

Microbial cultures are medically important for the isolation, detection, and differentiation of bacteria and other microbes that cause infectious disease. Characterization and identification using cultivation methods rely on both microbial phenotype and genotype. Cells can be isolated through inoculation of a streak plate to produce pure colonies. Selective media are used to inhibit the growth of certain groups of microbes, allowing others to grow. Differential media contain compounds that enable microbes to be distinguished visually by colony appearance or changes in surrounding media resulting from differences in metabolic or hemolytic activity. These media aid in the identification of bacteria and other microbes through differentiation and elimination.