- This involves taking DNA from two different sources and combining that DNA into a single molecule.
- The artificially created DNA is then reproduced by cloning.
- Six steps are involved in the process:
- Cutting (cleavage)
This involves cleavage of DNA to generate fragments of defined length, or with specific endpoints using restriction enzymes (see above). The DNA fragment of interest is called insert DNA.
1. Joining (ligation of DNA fragments)
A recombinant DNA molecule is usually formed by cleaving the DNA of interest to yield insert DNA and then ligating the insert DNA to vector DNA (recombinant DNA or chimeric DNA). DNA fragments are usually joined using DNA ligase.
The recombinant DNA molecule is introduced into a compatible host cell where it can replicate. The direct uptake of foreign DNA by a host cell is called genetic transformation. Recombinant DNA can be packaged into virus particles and transferred to host cells by transfection.
Cloning vectors allow replication and expression of recombinant DNA in host cells.
This involves the identification of host cells that contain the recombinant DNA of interest.