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A specialized transducing phage constructed from Bacillus subtilis phage phi 105.

T Iijima, F Kawamura, H Saito

    Gene
    |April 1, 1980
    PubMed
    Summary

    Researchers created a new defective phage, phi 105dmetB, by integrating Bacillus subtilis DNA into phage phi 105 DNA. This phage can transfer the metB+ gene, enabling genetic studies in Bacillus subtilis.

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    Area of Science:

    • Molecular Biology
    • Microbial Genetics
    • Bacteriophage Research

    Background:

    • Bacteriophages are viruses that infect bacteria and can be used as tools in genetic manipulation.
    • Bacillus subtilis is a model organism for microbial genetics research.

    Purpose of the Study:

    • To construct a new defective phage carrying a specific gene from Bacillus subtilis.
    • To characterize the genetic and physical properties of the newly constructed phage.
    • To investigate gene transfer mechanisms using bacteriophages.

    Main Methods:

    • Restriction endonuclease digestion (EcoRI) and in vitro ligation of phage and bacterial DNA.
    • Transformation of competent Bacillus subtilis cells.
    • Mitomycin C induction of prophages and subsequent transduction.

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  • Agarose gel electrophoresis and electron microscopy for physical mapping.
  • Transformation experiments to locate the metB+ gene.

    • Main Results:

    • A defective phage, phi 105dmetB, was successfully constructed by integrating Bacillus subtilis DNA into phage phi 105.
    • The new phage was capable of transducing the metB+ gene in Bacillus subtilis.
    • Physical mapping revealed that a foreign DNA fragment (EcoRI-M) replaced two adjacent fragments (EcoRI-G and E) in the phi 105 genome.
    • The metB+ gene was localized to the EcoRI-M fragment, which contained a BamHI-sensitive site.

    Conclusions:

    • The study successfully engineered a novel defective phage for gene transfer in Bacillus subtilis.
    • The metB+ gene is located on a specific foreign DNA fragment within the engineered phage genome.
    • This work provides a new tool for genetic analysis and manipulation of Bacillus subtilis.