Complementary DNA

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In genetics, complementary DNA (cDNA) is DNA synthesized from a mature mRNA template in a reaction catalyzed by the enzyme reverse transcriptase and the enzyme DNA polymerase.[1] cDNA is often used to clone eukaryotic genes in prokaryotes. When scientists want to express a specific protein in a cell that does not normally express that protein (i.e., heterologous expression), they will transfer the cDNA that codes for the protein to the recipient cell. cDNA is also produced by retroviruses (such as HIV-1, HIV-2, Simian Immunodeficiency Virus, etc.) which is integrated into its host to create a provirus.

Contents

Overview

The central dogma of molecular biology outlines that in synthesizing proteins, DNA is transcribed into mRNA, which is translated into protein. One difference between eukaryotic and prokaryotic genes is that eukaryotic genes can contain introns (intervening sequences), which are not coding sequences (exons), and must be removed from the RNA primary transcript before it becomes mRNA and can be translated into protein. Prokaryotic genes have no introns, so their RNA is not subject to splicing.

Often it is desirable to express eukaryotic genes in prokaryotic cells. A simplified method of doing so would include the addition of eukaryotic DNA to a vector, some times a prokaryotic host which would transcribe the DNA to mRNA and then translate it to protein. However, as eukaryotic DNA has introns, and since prokaryotes lack the machinery to splice them, the splicing of eukaryotic DNA must be done prior to adding the eukaryotic DNA into the host. This DNA, which was made as a complementary copy of the RNA and has no introns, is called complementary DNA (cDNA). To obtain expression of the protein encoded by the eukaryotic cDNA, prokaryotic regulatory sequences would also be required (e.g. a promoter).

Synthesis

Though there are several methods for doing so, cDNA is most often synthesized from mature (fully spliced) mRNA using the enzyme reverse transcriptase. This enzyme operates on a single strand of mRNA, generating its complementary DNA based on the pairing of RNA base pairs (A, U, G and C) to their DNA complements (T, A, C and G respectively).

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