cDNA library

A cDNA library is a collection of Complementary DNA copies made from an RNA population and retained for analysis, commonly as cloned inserts. It samples the transcripts present in the chosen material under the chosen preparation conditions. It is not a complete inventory of an organism's genome.

In a typical mRNA-derived preparation, reverse transcriptase makes the first DNA strand using RNA as template, a complementary strand is synthesized, and DNA copies are introduced into a cloning system. Mature processed transcripts can yield copies with joined exons rather than the original intervening introns. The representation can also depend on tissue, physiological state, RNA integrity, priming and cloning biases. Not every copy must be full length, and not every RNA-derived library is restricted to protein-coding messages.

A primary method by U. Gubler and B. J. Hoffman combined first-strand synthesis with RNase H/DNA-polymerase-I-mediated second-strand synthesis. They used sequencing of a known bovine preproenkephalin mRNA-derived product to test faithful full-length copying, rather than assuming that every recovered insert was complete.^[ref]

An original sampling calculation explains a limitation. If an independently sampled clone has probability $p$ of representing a target transcript, a library of $N$ clones contains at least one target with probabilityAt $p=10^{-4}$, $N=10,000$ gives about $0.632$, while $N=30,000$ gives about $0.950$. These are stipulated equal-sampling conditions; amplification and cloning biases can change the effective probability. Failure to find a clone does not establish absence of the gene or even absence of its transcript in the starting material.

A library can be screened by sequence complementarity or by a function expressed from its inserts. Those screens ask different questions: sequence recognition can recover a related fragment, while functional expression additionally requires a suitable insert and expression context. The sampling model, insert completeness and readout therefore belong to the interpretation of a negative screen.