Genetic Manipulation of Lytic Bacteriophages with BRED: Bacteriophage Recombineering of Electroporated DNA

We describe a recombineering-based method for the genetic manipulation of lytically replicating bacteriophages, focusing on mycobacteriophages. The approach utilizes recombineering-proficient strains of Mycobacterium smegmatis and employs a cotransformation strategy with purified phage genomic DNA a...

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Detalles Bibliográficos
Autores principales: Marinelli, L.J., Piuri, M., Hatfull, G.F.
Formato: SER
Materias:
BRED
Electroporation
Mycobacteria
Mycobacteriophage
Recombineering
bacteriophage DNA
genomic DNA
oligonucleotide
Rac protein
recombinant protein
RecT protein
bacteriophage recombineering of electroporated DNA
bacterium culture
electroporation
genetic engineering
genetic manipulation
genetic recombination
methodology
mutant
mycobacteriophage
Mycobacterium smegmatis
natural science
nonhuman
polymerase chain reaction
prophage
recombineering
Acceso en línea:http://hdl.handle.net/20.500.12110/paper_10643745_v1898_n_p69_Marinelli
Aporte de:
Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) de Universidad de Buenos Aires
Descripción
Sumario:We describe a recombineering-based method for the genetic manipulation of lytically replicating bacteriophages, focusing on mycobacteriophages. The approach utilizes recombineering-proficient strains of Mycobacterium smegmatis and employs a cotransformation strategy with purified phage genomic DNA and a mutagenic substrate, which selects for only those cells that are competent to take up DNA. The cotransformation method, combined with the high rates of recombination obtained in M. smegmatis recombineering strains, allows for the efficient and rapid generation of bacteriophage mutants. © 2019, Springer Science+Business Media, LLC, part of Springer Nature.

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