Complementary DNA

Patentability of cDNA was a subject of a 2013 US Supreme Court decision in Association for Molecular Pathology v. Myriad Genetics, Inc. As a compromise, the Court declared, that exons-only cDNA is patent-eligible, whereas isolated sequences of naturally occurring DNA comprising introns are not.

[3] In cellular life, cDNA is generated by viruses and retrotransposons for integration of RNA into target genomic DNA.

In molecular biology, RNA is purified from source material after genomic DNA, proteins and other cellular components are removed.

[7] Importantly, RNA integrity is maintained by inactivating RNases with chaotropic agents such as guanidinium isothiocyanate, sodium dodecyl sulphate (SDS), phenol or chloroform.

Various commercial kits exist for simple and rapid RNA extractions for specific applications.

The M-MLV reverse transcriptase from the Moloney murine leukemia virus is commonly used due to its reduced RNase H activity suited for transcription of longer RNAs.

[11] The AMV reverse transcriptase from the avian myeloblastosis virus may also be used for RNA templates with strong secondary structures (i.e. high melting temperature).

[12] cDNA is commonly generated from mRNA for gene expression analyses such as RT-qPCR and RNA-seq.

An optimized mixture of oligo-dT and random hexamer primers increases the chance of obtaining full-length cDNA while reducing 5' or 3' bias.

With amplification of DNA sequences via polymerase chain reaction (PCR) now commonplace, one will typically conduct reverse transcription as an initial step, followed by PCR to obtain an exact sequence of cDNA for intra-cellular expression.

This is achieved by designing sequence-specific DNA primers that hybridize to the 5' and 3' ends of a cDNA region coding for a protein.

They typically also contain a strong promoter to drive transcription of the target cDNA into mRNA, which is then translated into protein.