How is recombinant DNA made?

Recombinant DNA is an artificial DNA molecule created by combining DNA fragments from at least two different sources that are not necessarily related to one another. Recombinant DNA technology is based on introducing and incorporating a genetically modified vector into the genome of the host, which alters the phenotype of the host. 

There are 4 main steps involved in making recombinant DNA.

1. DNA Isolation - DNA is enclosed within the nucleus and typically coexists with other macromolecules such as proteins, RNA, lipids, and polysaccharides. To make recombinant DNA, the DNA must first be isolated (separated and purified) to obtain it in its purest form without any other macromolecules. Isolating DNA can be done using enzymes such as ribonuclease, lysozymes, chinitase, proteases, and cellulase. 
2. DNA Splicing or Fragmentation - The isolated and purified DNA is treated with restriction endonucleases, which splice the DNA into fragments. Restriction endonucleases are sequence-specific. They examine the length of DNA and splice it at specific sites called the restriction site. This helps to identify the exact site where the desired gene is inserted into the vector genome. 
3. DNA Amplification - Once the gene of interest has been spliced using the restriction enzymes, polymerase chain reaction (PCR) is used to amplify copies of the gene. PCR involves three stages - denaturation, annealing and extension. This process can create several thousand copies of DNA from a single copy or a few copies of DNA.  
4. DNA Ligation - The enzyme DNA ligase is used to link the vector of interest and the purified DNA fragment. The DNA molecule that results from the linkage is a hybrid of two DNA molecules – the vector and the interest molecule. In the terminology of genetics, this merging of distinct DNA strands is called recombination and the new hybrid DNA molecule is referred to as a recombinant DNA molecule. The recombinant DNA (rDNA) manufactured by the above process is transported into a recipient host cell, usually a bacterial cell, through a process known as transformation.