Gene Libraries with Big Pieces

A third type of gene library we will briefly describe has clones of large segments of the chromosome that could contain many genes. The motive behind making these kinds of libraries is that through gene mapping, we often know the position of a gene and it’s closest DNA neighbors. If we clone a piece of the chromosome that contains the gene of interest and the neighboring genes, gene mapping knowledge can be used to isolate a clone of that gene.

These libraries are made by combining the genomic DNA from an organism with vectors that make either yeast artificial chromosomes (YACs) or bacteria artificial chromosomes (BACs). YACs and BACs are recombinant DNA but because they are so large, they are more like chromosomes than plasmids. Yacs are transformed into yeast and BACs are transformed into bacteria. The geneticists generate large chunks of the chromosome by cutting carefully isolated chromosomes with restriction enzymes that recognize eight nucleotide pair sequences that rarely occur on the chromosome.

Fig.16: Genes that have a well mapped position on a chromosome using molecular (DNA) markers can be cloned by 'map based cloning'. (Image by D. Lee)


Fig.17: Large chromosome segments are combined with bacteria DNA to make artificial chromosomes (BACs). BACs with the gene of interest will also have DNA markers that map very close to this gene.  (Image by D. Lee)

A BAC or YAC that has the gene of interest can be identified from the library by screening with DNA markers. If the DNA marker maps near the gene, then a large insert identified that has DNA markers mapping to both sides of the gene should have the gene of interest as well.

Geneticists have many tools and strategies at their disposal to make gene libraries. Just a few have been described here. Once the gene library is made, the geneticist must then have a plan for identifying the bacteria colony that is cloning the gene they are interested in. This final stage of gene cloning is called library screening and is described in the next lesson.