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Deletion between direct repeats in bacteriophage T7 gene 1.2
1Temple University School of Medicine, Department of Biochemistry and Molecular Biology, Philadelphia, PA 19140.
Mutation Research
|August 1, 1993
Summary
Bacteriophage T7 deletion mutagenesis was studied using foreign DNA inserts. Deletion rates were similar across different gene locations and unaffected by T7 ligase at 37°C, but influenced by DNA Polymerase I.
Area of Science:
- Molecular Biology
- Genetics
- Virology
Background:
- Bacteriophage T7 serves as a model organism for studying DNA replication and repair mechanisms.
- Understanding deletion mutagenesis is crucial for comprehending genome stability and evolution in viruses.
Purpose of the Study:
- To investigate deletion mutagenesis in bacteriophage T7 using insertion-reversion assays.
- To quantify deletion rates of foreign DNA inserts in different T7 genes.
- To assess the influence of T7 ligase and E. coli DNA Polymerase I on deletion rates.
Main Methods:
- Insertion-reversion assay with 10-bp direct repeats of foreign DNA in bacteriophage T7.
- Quantification of deletion rates using Luria and Delbrück fluctuation tests.
- Assaying phage growth on selection strains of E. coli to measure functional gene restoration.
Main Results:
- Deletion rates were consistent for inserts in both gene 1.2 and gene 1.3, indicating site and repeat sequence independence.
- Deletion was independent of functional T7 ligase at 37°C but increased in ligase-deficient phage at 43°C.
- E. coli DNA Polymerase I significantly decreased deletion rates, with polA1 mutation reducing it by 60% and polA546ex by 90%.
Conclusions:
- Deletion mutagenesis in bacteriophage T7 is not site-specific or dependent on the direct repeat sequence.
- T7 ligase plays a role in deletion at elevated temperatures, while E. coli DNA Polymerase I acts as a suppressor of deletion events.