This study is designed to elucidate the relationship between the structure of transposable elements and the mechanisms by which they join unrelated segments of DNA. We are investigating four systems: (1) lambda::Tn9. Lambda phages containing the chloramphenicol resistance element Tn9 will insert into the bacterial chromosome in the absence of the normal viral integration system. Previous results showed that insertion and excision of lambda::Tn9 prophages are precise events involving recombination at ISI elements at the prophage ends. We will examine the products of prophage excision to determine the nature of the excision reaction. (2) Artificial transposons constructed by recombination. A galE gene flanked by two ISI elements is transposable. The two ISI elements can be in parallel or opposed orientation. We will study the behavior of ISI-galE-ISI transposons to determine the importance of direct versus opposed terminal repeats on insertion and excision events. (3) Tn5. Substitution of trp DNA for certain parts of the Tn5 kanamycin-resistance transposon results in a recessive transposition defect. We will use in vivo and in vitro Tn5 mutants to determine what parts of the transposon encode diffusible activities required for transposition. (4) Tn3. Deletion of different regions of the Tn3 beta-lactamase transposon alter the structure of recombinant products (cointegration vs. transposition) catalyzed by the tnpA transposase gene product (TnpA activity). We will study the products of lambda and plasmid recombination catalyzed by the TnpA activity and various mutants of Tn3.
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