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A phasmid shuttle vector for the cloning of complex operons in Salmonella
L Gutmann1, M Agarwal, M Arthur
1Maxwell Finland Laboratory For Infectious Diseases, Boston University, School of Medicine, Massachusetts 02118.
Plasmid
|January 1, 1990
Summary
Phasmid P4 vir1 functions as a versatile cloning vector in Salmonella and E. coli. This phage shuttle vector enables efficient gene transfer, aiding in the study of bacterial virulence factors.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Phasmid P4 vir1 propagates in Escherichia coli as a lytic phage, plasmid, or prophage.
- P4 derivatives have been engineered as cloning vectors based on their propagation modes.
Purpose of the Study:
- To evaluate phasmid P4 as a cloning vector in Salmonella species.
- To demonstrate its utility as a high-efficiency phage shuttle vector for gene transfer between Salmonella and E. coli.
Main Methods:
- Propagation of P4 vir1 in Salmonella spp. and E. coli.
- Assessment of P4-mediated gene transfer efficiency.
- Cloning and expression of virulence factor operons (P pilus and hemolysin) from uropathogenic E. coli.
Main Results:
- Phasmid P4 propagates efficiently as a helper-dependent lytic phage and plasmid in Salmonella spp.
- P4 phage-mediated gene transfer in both E. coli and Salmonella spp. shows high efficiency, only 10-fold lower than plaquing efficiency.
- Gene transfer in Salmonella spp. is approximately 10^4-fold more efficient than conventional DNA transformation.
Conclusions:
- Phasmid P4 serves as a highly efficient phage shuttle vector for reversible gene transfer between Salmonella and E. coli.
- This system is valuable for analyzing pathogenic virulence factors, as demonstrated by cloning and expression of key operons.