Biotechnology is the use of biology to solve problems and make useful products. The most prominent area of biotechnology is the production of therapeutic proteins and other drugs through genetic engineering. The term ‘Biotechnology’ was coined in the year 1919 by an agricultural engineer Karoly Ereky, hence he is called as the father of Biotechnology.
The two core techniques that enabled birth of modern biotechnology are :
The techniques of genetic engineering mainly include:
The enzymes which include the restriction enzymes help to cut, the polymerases- help to synthesize and the ligases- help to bind. The restriction enzymes used in recombinant DNA technology play a major role in determining the location at which the desired gene is inserted into the vector genome. They are two types, namely Endonucleases and Exonucleases.
The Endonucleases cut within the DNA strand whereas the Exonucleases remove the nucleotides from the ends of the strands. The restriction endonucleases are sequence-specific which are usually palindrome sequences and cut the DNA at specific points.
They scrutinize the length of DNA and make the cut at the specific site called the restriction site. This gives rise to sticky ends in the sequence. The desired genes and the vectors are cut by the same restriction enzymes to obtain the complementary sticky notes, thus making the work of the ligases easy to bind the desired gene to the vector.
The vectors – help in carrying and integrating the desired gene. These form a very important part of the tools of recombinant DNA technology as they are the ultimate vehicles that carry forward the desired gene into the host organism. Plasmids and bacteriophages are the most common vectors in recombinant DNA technology that are used as they have a very high copy number. The vectors are made up of an origin of replication- This is a sequence of nucleotide from where the replication starts, a selectable marker – constitute genes which show resistance to certain antibiotics like ampicillin; and cloning sites – the sites recognized by the restriction enzymes where desired DNAs are inserted.
Host organism – into which the recombinant DNA is introduced. The host is the ultimate tool of recombinant DNA technology which takes in the vector engineered with the desired DNA with the help of the enzymes.
There are a number of ways in which these recombinant DNAs are inserted into the host, namely – microinjection, biolistics or gene gun, alternate cooling and heating, use of calcium ions, etc.
Recombinant DNA Technology is the technology to produce an artificial DNA molecule by combining two or more fragments of DNA which are not necessarily associated with each other. Usually, such DNA fragments are obtained from several biological sources.
DNA isolation: DNA is isolated in its pure form, which means they are devoid of other macromolecules.
Cutting of DNA: For this step, the restriction enzymes are quite vital. It helps to identify the location wherein a designated gene is introduced into a vector genome. The said reaction is known as restriction enzyme digestions.
Amplifying of DNA: Copies of genes are amplified through PCR or polymerase chain reaction. It is essentially a process to increase a single DNA copy into several copies after the desired gene of interest is cut with restriction enzymes.
Joining DNA: The vector and a section of DNA is joined in this step. It is achieved with the help of enzyme DNA ligase.
Insertion of rDNA into a host: Here rDNA is added to the recipient host cell, and the entire process is called transformation. Post insertion, the recombinant DNA multiplies and manifests as manufactured protein under favourable conditions.