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Transposition of mini-Mu containing only one of the ends of bacteriophage Mu
Abstract:
Transposition of mini-Mu containing only one of the ends of bacteriophage Mu was studied. Transposition was observed when plasmids containing this mini-Mu were used as the donor in a mating-out assay with the F factor POX38, which lacks all known transposable elements, as the target. Analysis of the plasmids isolated from the recipient strain showed that in most cases the mini-Mu containing plasmid and POX38 were fused and that a part of the plasmid had been duplicated, indicating the formation of co-integrates. To obtain the DNA sequences of the junctions between donor and recipient DNA, an F factor was constructed that made it possible to analyse these junctions directly. The results showed that several sequences can be used as an alternative end in transposition and that these alternative ends show homology with the consensus for a strong A binding site. Moreover, the first base pair at the junction was always a (TA) base pair. This base pair is situated at exactly the same position with respect to the sequence which has homology with the consensus for a strong A binding site as in the ends of Mu.
Insights
This study reveals that mini-Mu DNA can transpose using alternative DNA sequences as ends, forming fused DNA structures. These alternative ends share similarities with bacteriophage Mu ends, influencing transposition outcomes.
Area of Science:
- Molecular Biology
- Genetics
- Microbiology
Background:
- Bacteriophage Mu is a well-characterized transposable element.
- Understanding transposition mechanisms is crucial for genetic engineering and genome stability.
- Mini-Mu elements, derived from bacteriophage Mu, retain transposition capabilities.
Purpose of the Study:
- To investigate the transposition of mini-Mu elements lacking one native end.
- To identify alternative DNA sequences that can function as transposition ends.
- To characterize the DNA junctions formed during mini-Mu transposition.
Main Methods:
- Utilizing a mating-out assay with a mini-Mu containing plasmid as donor and F factor POX38 as target.
- Analyzing plasmids from recipient strains to identify co-integrate formation.
- Constructing a specialized F factor for direct analysis of DNA junction sequences.
Main Results:
- Mini-Mu transposition was observed, leading to the formation of co-integrates between the donor plasmid and the target F factor.
- Several DNA sequences, distinct from native Mu ends, were identified as functional alternative transposition ends.
- These alternative ends exhibited homology to a strong A-binding site consensus sequence.
- Transposition junctions consistently initiated with a (TA) base pair, positioned similarly to native Mu ends relative to the homology site.
Conclusions:
- Mini-Mu transposition is not strictly dependent on its native ends and can utilize alternative DNA sequences.
- The identified alternative ends share functional and sequence similarities with native Mu ends, suggesting a conserved recognition mechanism.
- The consistent (TA) base pair at the junction provides insights into the precise initiation of transposition by mini-Mu.