Preparation of BAC Libraries From Bacterial Genomes by In Vitro Packaging
Genomic libraries represent a powerful resource for genetic studies of bacteria. Large-insert libraries in bacterial artificial chromosome (BAC) vectors are particularly important not only for genome-sequencing projects, but for comparative and functional genomic studies once a complete sequence is known. Cloned DNA can be introduced into bacterial cells in two different ways. One way is to have cells take up naked DNA; this is known as transformation (1). The other way is to package the recombinant DNA inside bacteriophage particles in vitro using a phage such as λ. This process, known as in vitro packaging, allows DNA to be introduced by infection (2). Although transformation allows DNA of any size to be introduced, DNA uptake occurs at very low frequency. On the other hand, in vitro packaging limits the size of the DNA based on the phage head stability. However, in vitro package is the most efficient method for introducing large-insert clones.
KeywordsSucrose Glycerol EDTA MgSO4 Isopropanol
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