Selection of Recombinant Vector

It may be pointed out that when an experiment is performed to insert a DNA fragment into a
vector, two types of vectors molecules are obtained (1) many vector molecules will contain the DNA insert but, (2) many others well contain only the vector sequences. This mixture of vector molecules is used for transformation of host cell. (1) Some host cells will receive the recombinant vector, (2) some others will contain the normal unaltered vector, while (3) the majority of them will contain no vector, i.e will not be transformed. In a cloning experiment it is critical to effectively select for the low frequency of cells transformed by the recombinant vector form among the cells containing the un altered vector and the non transformed cells.
Selection of host cells transformed by the recombinant vector is easily achieved by placing two selectable markers, e.g. antibiotics resistance gens, such as a ampicillin resistance and tetracycline
resistance in the vector. The DNA insert is integrated within one of the two selectable rmarkers. If the DNA insert is integrated with in the ampicillin resistance gene, the cells containing recombinant vector will be resistant to tetracycline but sensitive to amipicillin. In contrast, non transformed cells will be sensitive to both the antibiotics, while that containing the unaltered vector will is resistant to both. Therefore, following transformation with the above recombinant vectors cells are plated on a tetracycline supplemented medium this eliminates the nontransformed cells. The remaining colonies are not replica plated on ampicillin vector, and are isolated from the master plate. Further, transformed cells tend to lost the recombinant vector, since cells lacking such vectors divide much faster. The use of a vector having two selectable markers allow the maintenance of cells containing the recombinant vector on antibiotic medium which eliminates the vector free cells produced during culture.