A coliphage lambda vector with enhanced biological containment: lambda gtALO.lambda B.

Abstract

The biological containment of the lambda gt family of cloning vectors has been enhanced by conditionally blocking DNA replication as well as head and tail morphogenesis. The vector, lambda gtALO.lambda B, was constructed by crossing the Oam29, Aama1 and Lam439 mutations into lambda gt.lambda B. The mutation blocking phage DNA replication, Oam29, is suppressed by suII+ or suIII+. The head gene mutation, Aama1, is suppressed by suIII+ but not by suII+ and the tail gene mutation, Lam439, is suppressed by suII+ but not by suIII+. This allows the option of increasing the biological containment by producing heads when a large amount of cloned DNA is being prepared from an individual isolate. A model recombinant, lambda gt Aama1 Lam439 Oam29.KmR' (lambda gtALO.KmR') was constructed and the containment of the vector was evaluated by the series of standardized experiments required for EK2 certification.