Selectable Marker

Introduction of plasmids in to E. coli cells is an inefficient process. Thus, a method of selecting those cells that have received a plasmid is required. Furthermore, cells that do not contain a plasmid are at a growth advantage over those that do and, thus, have to replicate both the chromosome and additional plasmid DNA. This is of par­ticular consequence when dealing with high-copy-number or large plasmids. In this case, a selective pressure must be imposed for maintenance of the plasmid. Almost all conventional plasmids use an antibiotic resistance gene as a selectable marker, carried on the backbone of the vector. Thus, the addition of the appropriate antibiotic to the growth medium will kill those cells that do not contain the plasmid and produce a culture in which all cells do contain a plasmid. In many cases, the choice of antibiotic is not restricted. However, some cloning strains of E. coli are inherently resistant to some antibiotics and, thus, the same antibiotic cannot be used as a selection for those cells carrying a particular plasmid. The genotype of the desired cloning strain should be checked prior to cloning (see Part 3). In some situations, downstream applica­tions render some antibiotics as unsuitable choices