The aim of this work was to develop a more efficient and usable basic form of cloning vehicle for genetic engineering than those currently in use. Subsequent work would then enable a new generation of cloning vectors to be constructed. The characteristics that such a vehicle would require were determined by considering the basic processes involved in DNA cloning and the kinds of facilities that vector replicas can provide. Against this background the development of currently used phage and plasmid vectors was considered and the properties they offer examined. Both growth forms have different kinds of advantages and disadvantages in use. It is postulated that a plasmid/phage growth cycle of the vector replicon would harness the advantages of both growth phases and enable their disadvantages to be avoided. Consequently, a plasmid-phase growth cycle of bacteriophage lambda has been developed to fulfil this requirement. Immediately after infection of a bacterial cell,the phage lambda genome exists transiently as a plasmid replicon before growth diverges to the lysogenic or lytic pathways. A potential therefore exists for the development of an alternative plasmid growth pathway. Evidence had suggested that an N mutation arrests development at this stage (Signer, 1969;Lieb, 1970;Kleckner and Signer, 1977). However,this investigation found that such plasmid formation is abortive. A series of further experimental investigations is reported that have enabled the elucidation of the genetic elements controlling the plasmid pathway. The plasmid mode of growth of lambda genomes is subject to a novel kind of fine genetical control, quite different to that of the lytic or lysogenic modes. Furthermore, lambda genomes growing as plasmids can be recovered as phages, if subjected to a precise molecular switch:-this facilitates the PHASMID growth cycle. Application of lambda phasmids for DNA cloning is considered. Precise genetical control and characterization,coupled with possible alternate growth phases, make lambda phasmids potentially extremely useful for the propagation and manipulation of cloned genes.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:674130 |
| Date | January 1980 |
| Creators | Hadfield, Christopher |
| Publisher | University of Leicester |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/2381/35238 |
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