Origin of replication (ori)
• This is a DNA sequence that is responsible for initiating replication. Any piece of DNA when linked to this sequence can replicate within the host cells.
• ori also controls the copy numbers of the linked DNA.
Selectable marker
• It helps to select the host cells which contain the vector (transformants) and eliminate the non-transformants.
• Transformation is defined as the procedure by which a piece of DNA is introduced into a bacterial host.
• Genes encoding resistance to antibiotics like ampicillin, chloramphenicol, tetracycline or kanamycin, are useful selectable markers for E. coli.
• The normal E. coli cells do not carry resistance against any of these antibiotics.
Cloning sites
• To link the alien DNA, the vectors require very few (mostly single) recognition sites for the restriction enzymes.
• More than one recognition sites within the vector, can complicate the gene cloning as it will generate several fragments.
• Ligation of alien DNA can be carried out at a restriction site present in one of the two antibiotic resistance genes.
