An expression-packaging-processing vector which selects and maintains 7-kb DNA inserts in the blue T4 phage genome.
We have developed an efficient positive-selection vector to insert foreign DNA segments fused to the T4 ipIII gene (encoding internal protein IPIII) into the bacteriophage T4 genome. By using partial deletions of the T4 e gene, which encodes phage lysozyme, lysozyme activity required for plaque formation is used to select plasmid integrants which restore the e gene. In this work, we demonstrate that DNA inserts more than 7.0 kb in length can be incorporated into a T4 genome lacking the alt gene. In addition, the recombinant T4 not only contains a fusion gene driven by the T4 ipIII promoters, but also packages the fusion protein into the T4 capsid due to targeting by the IPIII portion. This expression-packaging-processing system shows that active IPIII::beta Gal fusion reporter protein is produced and packaged during phage infection.
Hong
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YR
Department of Biological Chemistry, University of Maryland Medical School, Baltimore 21201-1503.
Black
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LW
D017122:Bacteriophage T4 / Q000235:genetics*; D015139:Blotting, Southern; D002213:Capsid / Q000235:genetics; D036022:Capsid Proteins; D005814:Genes, Viral; D005822:Genetic Vectors; D007763:Lac Operon; D009113:Muramidase / Q000235:genetics; D010641:Phenotype; D011995:Recombination, Genetic; D015183:Restriction Mapping
D036022:Capsid Proteins; C081364:internal protein III, bacteriophage T4; D009113:Muramidase