A genomic library is a comprehensive collection of cloned DNA fragments derived from a genome. Each part of the genome is represented in the library several times, and the number of times it is represented on average is called the coverage of a library. The library can be screened for the presence of the sequence of interest by radioactively labelling the DNA (usually between 100 and 500 nucleotides long) and using this as a probe to identify the clone that contains the selected sequence. The clone selected can then be grown in bacteria to produce large amounts of clone DNA, which can be studied. If for example, the sequence of interest was part of a gene, by using this sequence as a probe, the clone containing, hopefully, the whole gene could be isolated.
The DNA fragments are generated from genomic DNA by treatment by restriction enzymes--enzymes that cut at specific sites in the DNA. The genomic DNA is only partially cut with these enzymes; not all sites are cut, this is termed a restriction enzyme partial digest. The size of fragments is determined by the type of restriction enzyme used. For example, if the DNA is cut with an enzyme that cuts at the sequence GATC, then the fragments produced will be small because there is a GATC site about every 275 nucleotides in DNA. However, an enzyme that cuts at the sequence GCGGCCGC produces larger fragments, because this sequence is less common in the genome. The DNA fragments can be cloned into different vectors depending on the size of the fragment. For fragments of up to 10kb, plasmids are suitable vectors; cosmids can carry up to 50kb, bacterial artificial chromosomes up to 300kb and yeast artificial chromosomes (YACs) up to 2Mb. As their name suggests, YACs are grown in yeast cells rather than than the bacteria E.coli.
The clone DNA from the thousands of clones produced are then usually placed as a grid pattern on a nylon filter (gridded filter) by a robot. This nylon filter, after being treated with ultraviolet light to fix the DNA onto it, can be screened with the radioactively labelled probe of choice. Screening is performed by DNA hybridization. The probe hybridizes to the clone DNA that contains the sequence. The coordinates of the clone on the gridded filter can then be read and the appropriate clone DNA grown and purified.
Genomic libraries have been used in human genetics as part of the human genome project (HGP). Whole genome BAC libraries and libraries made from single flow-sorted chromosomes were used. The RPCI-11 library is a typical genomic BAC library and was used extensively by the HGP, and has 32x coverage of the genome. Genomic BAC libraries of several other organisms have been produced, such as chimpanzee, baboon, rat, mouse, fruit fly and zebrafish. As well as enabling large physical maps to be created, these gridded filters were copied, and are made available to independent scientists. Once the independent scientist has identified the clone of interest, he or she can contact the laboratory that produced the library and they can provide a bacterial colony containing the clone DNA. This means that a collection of genomic libraries can be used by many people around the world preventing duplication of effort by creating further genomic libraries.
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