Re: What is a cDNA library?

Before you can understand what a cDNA library is, you need to understand two things: what is cDNA; and what do molecular biologists mean by a "library".

The genetic material of the cell is composed of Nucleic Acids. These can be separated into two forms: deoxyribo-nucleic acids (DNA) which make up the chromosomes; and ribo-n ucleic acids (RNA) which decode the genes encoded in the chromosomal DNA and use the information to produce proteins for the cell. When a gene is activated (i.e. made available for usage), an enzyme called RNA polymerase makes an RNA copy of the gene (called an hnRNA; hn is for heavy, nuclear), which is then processed into a more compact form (called mRNA; m is for messenger) that exits the nucleus and is used as a template for protein production. One of the major differences between hnRNA and mRNA is the existence of introns. Introns are present in chromosomes as non-coding stretches of DNA which break up individual genes into small, separated fragments, called exons. When RNA polymerase transcribes a gene, it copies the introns and exons together, so that the resulting hnRNA contains the fragmented gene plus all of its introns. A group of RNA-protein enzymes (called snRNP's) attach to the introns in hnRNA's to form Spliceosomes, which excise the introns and splice the exons together to form the entire, uninterrupted gene. After other modifications, the result is an intronless mRNA copy of the gene.

The only problem with mRNA is that, for various reasons, it is much more difficult to work with, in the laboratory, than DNA. Fortunately, all RNA viruses (including Poliovirus, Herpesvirus, HIV, and many more) produce an enzyme called Reverse Transcriptase (RT) which makes DNA copies of RNA strands and is easy to mass produce from bacterial cultures. Because the DNA is a copy of an RNA, rather than vice versa, it is called cDNA (c is for copy). The most common usage of RT is to make cDNA from mRNA. cDNA has two advantages over chromosomal DNA: there are no introns, so it is easier to identify and characterize the genes; and cDNA only represents those genes that are being actively used by the cell, since RNA polymerase only transcribes activated genes.

Now for the "library". If you have a piece of a gene and you want the rest of the gene, it would take a very long time to search from one end of a genome to the other looking for your gene. On the other hand, if you divide the genome into fragments, and then identify which fragment contains your gene, it takes very little time to search from one end of a fragment to the other. This is essentially what libraries are about. To make a library, you divide a large pool of DNA into smaller units, and then give each unit the ability to replicate independently, by splicing it into a vector (like a virus or an artificial chromosome), and cloning it into a cell which will reproduce and make copies. Genomic libraries exist for all organisms commonly used in the lab, and consist of enzymatically digested chromosome fragments spliced into various vectors and placed in various cells depending on the size of the fragments (phage libraries in bacteria for small fragments to YAC libraries in yeast for huge fragments).

cDNA libraries are simpler to construct, because cDNA's, like their parental mRNA's, are already fairly short, so an entire cDNA can be spliced into a single vector. The reason you need to make a library is that cells produce tens of thousands of different mRNA's at a time, so that after using RT to make cDNA, you still have a massive pool of different cDNA's with which to work. As stated above, cDNA libraries have advantages over genomic libraries: there are no introns, so there is no danger of pieces of your gene being chopped onto separate clones; and the library is (hopefully) enriched for your gene, since instead of one or two copies, as in the genomic library, you have as many copies as the cell could produce mRNA's for that gene. So most molecular biologists, when searching for a new gene, start by screening a cDNA library from a tissue or organism that they suspect is actively using that gene. Most new genes are found this way.

As an aside, one of the post-doc in my lab says that a cDNA library is a room full of books about cloning. He's not allowed to answer anymore questions.