TOPO cloning is a molecular biology cloning technique in which DNA fragments are rapidly joined to plasmid vectors by the enzyme topoisomerase I, without the requirement for DNA ligase.
What are the types of TOPO cloning?
There are three types of TOPO cloning:
- Sticky-end TA cloning
- Blunt-end cloning
- Directional cloning
Sticky-end cloning and blunt-end cloning are bidirectional with inserts able to ligate in either orientation into the vector. Directional cloning ensures inserts ligate in a preferred orientation.
Sticky-end TOPO TA Cloning
Sticky-end TOPO cloning exploits the inherent activity of Taq polymerase, which adds a single deoxyadenosine (A) to the 3′-end of PCR products. Inserts are PCR amplified and then ligated into linearized TOPO TA vectors which have a complementary deoxythymidine (T) overhang.