Construction of Genomic Libraries in λ-Vectors

Abstract

Lambda (λ) bacteriophages are viruses that specifically infect bacteria. The genome of λ-phage is a double-stranded DNA molecule approx 50 kb in length (1). In bacterial cells, λ-phage employs one of two pathways of replication: lytic or lysogenic. Commonly, λ-phage vectors replicate via the lytic pathway. During lytic growth, the viral DNA is replicated manifold, a large number of phage gene products are synthesized, and progeny phage particles are assembled. The cell is eventually lysed, releasing its many new infectious virus particles; at this time, plaques will form on an infected bacterial lawn. Its high infectivity and clone-style propagation are the inherent basis for the use of λ-phage as a vector. Moreover, the central third of the viral genome is not essential for lytic growth and, thus, can be replaced by a variety of foreign gene segments. λ-Vectors usually contain multiple cloning sites facilitating the cloning of foreign DNA. In addition, λ-phage vectors have the following advantageous features: high cloning efficiency, a relatively large insert-size capacity, and suitability for screening using nucleic acid probes. In terms of gene screening, genomic DNA libraries are often screened by hybridization using a radioactive nucleic acid probe.