cDNA (Complementary DNA)
The term cDNA refers to complementary DNA. cDNA is known to be synthesized, or manufactured from an mRNA or messenger RNA template. It is synthesized in a reaction that is catalyzed by the reverse transcriptase and DNA polymerase enzymes. Essential to note is that cDNA is usually used to clone eukaryotic genes in prokaryotes.
Scientists usually use cDNA when they want to express s certain protein in a cell that does not normally express such a protein. This process is referred to as heterologous expression. The expression of such a protein will be done by transferring the cDNA that codes for that protein to the recipient cell. Also, essential to note is that cDNA can also be produced by retroviruses like Simian Immunodeficiency Virus, HIV-1 and HIV-2 among others. Once the cDNA is created from such viruses, it is integrated into the genome of the host, where it goes on to create a provirus.
Research shows that when a protein is being synthesized, a gene’s DNA is transcribed into an mRNA, which is then translated into a protein. Genes are usually divided into eukaryotic and prokaryotic genes. The only difference between these genes is that the eukaryotic genes contain introns instead of extrons that are contained in the prokaryotic genes.
Introns are not coding sequences, while extrons are DNA coding systems. During the transcription of the proteins, all intron RNA are cut from the primary RNA and the remaining piece is sliced back to become an mRNA. In other words, the mRNA is formed after all introns are removed from the primary RNA. Once formed, the mRNA is then translated into an amino acid and comprises a newly formed protein. From the above, it is noted that prokaryotic genes do not contain any introns, so their RNAs are not subject to cutting or splicing.
In some instances, it might be necessary to make a prokaryotic cell express the genes of a eukaryotic cell. One of the ways of making this possible is by adding eukaryotic DNA directly into the prokaryotic cell, so as to allow it to make a protein of its own. As has been noted, the eukaryotic DNA has introns, while the prokaryotic DNA does not have the machinery for removing introns from the RNA that has been transcribed. As a result, all intron sequences need to be removed from the eukaryotic DNA before it is transferred to the prokaryotic cell. This way, the cell will not be placed with the burden of having to remove introns. The intron-free DNA that is created is as a result of intron-free mRNA, which is why it is referred as a complementary copy of the mRNA. It is for this reason that is referred to as a complementary DNA or cDNA.
Though there are numerous processes for synthesizing cDNA, the best way of doing so is by using mature or fully spliced mRNA. This is usually done by using the enzyme reverse transcriptase. This enzyme is used because it mainly operates as a single strand of mRNA. It generates its cDNA by pairing RNA base pairs to their DNA complements.