Introduction of transposon Tn5 into Myxococcus for analysis of developmental and other nonselectable mutants

Abstract

The transposon Tn5, which carries a gene for kanamycin resistance, can be introduced into Myxococcus xanthus, an organism that undergoes a primitive cycle of development, from Escherichia coli by the specialized transducing phage P1::Tn5. Tn5 DNA sequences, but no P1 sequences, are found in the stable kanamycin-resistant transductants. Tn5 transposes from P1 to many different chromosomal sites in Myxococcus. In each independent transductant of Myxococcus examined, the Tn5 element is found in a different DNA fragment produced by cleaving cell DNA with a restriction endonuclease. Moreover, different Tn5 insertions have been found linked to the first 20 different genetic sites tested. Once inserted into the Myxococcus chromosome, Tn5 remains fixed in position during growth and when transferred to another strain by generalized transduction. To analyze developmental or other mutants that have no selectable phenotype themselves, a general method has been devised, and tested, for the systematic isolation of a Tn5 insertion near any arbitrary locus.