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Abstract:
To ascertain the form and cellular location of the copies of bacteriophage Mu DNA synthesized during lytic development, DNA from an Escherichia coli lysogen was isolated at intervals after induction of the Mu prophage. Host chromosomes were isolated as intact, folded nucleoids, which could be digested with ribonuclease or heated in the presence of sodium dodecyl sulfate to yield intact, unfolded nucleoid DNA. Almost all of the Mu DNA in induced cells was associated with the nucleoids until shortly before cell lysis, even after unfolding of the nucleoid structure. We suggest that the replicas of Mu DNA are integrated into the host chromosomes, possibly by concerted replication-integration events, and are accumulated there until packaged shortly before cell lysis. Nucleoids also were isolated from induced lambda lysogens and from cells containing plasmid DNA. Most of the plasmid DNA sedimented independently of the unfolded nucleoid DNA, whereas 50% or more of the lambda DNA from induced lysogens cosedimented with unfolded nucleoid DNA. Possible explanations for the association of extrachromosomal DNA with nucleoid DNA are discussed.
Insights
Newly synthesized bacteriophage Mu DNA associates with host cell nucleoids during lytic development. This suggests Mu DNA integrates into host chromosomes before packaging and cell lysis.
Area of Science:
- Molecular Biology
- Virology
- Microbiology
Background:
- Bacteriophage Mu is a temperate bacteriophage that infects Escherichia coli.
- Understanding the replication and integration of bacteriophage DNA is crucial for studying viral life cycles.
- The cellular location and form of viral DNA during lytic development are key aspects of viral replication.
Purpose of the Study:
- To determine the cellular location and physical state of bacteriophage Mu DNA during its lytic cycle in Escherichia coli.
- To investigate the association of newly synthesized Mu DNA with host cell structures.
Main Methods:
- Isolation of host nucleoids from Escherichia coli lysogens at various time points after Mu prophage induction.
- Treatment of nucleoids with RNase and SDS to unfold the structure.
- Sedimentation analysis to determine the association of Mu DNA with nucleoids.
- Comparison with DNA association in induced lambda lysogens and plasmid-containing cells.
Main Results:
- Newly replicated bacteriophage Mu DNA remains associated with host cell nucleoids throughout most of the lytic cycle, even after unfolding.
- This association persists until shortly before cell lysis.
- Extrachromosomal lambda DNA also shows association with nucleoids, while plasmid DNA largely sediments independently.
Conclusions:
- Bacteriophage Mu DNA replicas are likely integrated into host chromosomes during replication.
- Accumulation of Mu DNA within the host chromosome precedes its packaging for lysis.
- The association of extrachromosomal DNA with nucleoids warrants further investigation.