cDNA

Complementary DNA (cDNA) is a form of DNA synthesized from a single-stranded RNA (e.g., messenger RNA (mRNA)) template in a reaction catalyzed by the enzyme reverse transcriptase. cDNA is often used in genetic engineering, gene cloning, and PCR (Polymerase Chain Reaction) because it is composed only of the coding regions of a gene, excluding introns found in the pre-mRNA.

Overview

cDNA is generated by reverse transcribing mRNA. Since mRNA is processed and only includes the exons of a gene, the cDNA generated from it can be used to clone eukaryotic genes in prokaryotes. This is because prokaryotes, which do not have nuclear compartmentalization, cannot splice out introns as eukaryotic cells can.

Production

The production of cDNA involves the following steps:

1. Extraction of RNA from a cell
2. Using the enzyme reverse transcriptase to synthesize a DNA strand complementary to the mRNA template
3. Depending on the application, the single-stranded cDNA may be converted into double-stranded DNA (dsDNA) using DNA polymerase.

Applications

cDNA has several important applications in the field of biotechnology and genetics:

• Gene cloning and expression: cDNA is used to clone eukaryotic genes in prokaryotes. Since cDNA is devoid of introns, it can be expressed by prokaryotic cells, which cannot remove introns.
• PCR amplification: cDNA can serve as a template for PCR, a technique used to amplify specific DNA sequences.
• Gene expression studies: cDNA libraries can be used to study gene expression in various tissues and under different conditions.

cDNA Libraries

A cDNA library is a collection of cDNA clones that represent the mRNA transcripts present in a particular biological sample. These libraries are valuable resources for studying gene expression, identifying novel genes, and understanding genetic functions.

Challenges and Limitations

While cDNA is a powerful tool in genetic research, it has limitations:

• It only represents the exons of genes, so information about regulatory elements present in introns and non-coding regions is not included.
• The quality of cDNA depends on the integrity of the mRNA from which it is synthesized, which can be degraded during extraction.

See Also

• DNA
• RNA
• Gene expression
• Molecular cloning