Construction of a New Phage Integration Vector pFIV-Val for Use in Different Francisella Species

We recently identified and described a putative prophage on the genomic island FhaGI-1 located within the genome of AS02-814 ( subsp. -like 3523). In this study, we constructed two variants of a phage integration vector, called pFIV1-Val and pFIV2-Val ( Integration Vector-tRNA -specific), using the...

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Bibliographic Details
Published in:Frontiers in cellular and infection microbiology 2018-03, Vol.8, p.75-75
Main Authors: Tlapák, Hana, Köppen, Kristin, Rydzewski, Kerstin, Grunow, Roland, Heuner, Klaus
Format: Article
Language:English
Subjects:
Bacteriophages - genetics
Chloramphenicol Resistance - genetics
DNA, Bacterial - genetics
Drug Resistance, Bacterial - genetics
episomal
Escherichia coli - genetics
Francisella - genetics
Francisella - growth & development
Francisella - virology
Francisella tularensis
Francisella tularensis - genetics
Genetic Vectors
genomic island
Genomic Islands
Humans
Integrases - genetics
integrative vector
Microbiology
Mutation
pFIV-Val
phage
Plasmids
Recombination, Genetic
RNA, Transfer, Val - genetics
Transformation, Bacterial
U937 Cells
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Summary:We recently identified and described a putative prophage on the genomic island FhaGI-1 located within the genome of AS02-814 ( subsp. -like 3523). In this study, we constructed two variants of a phage integration vector, called pFIV1-Val and pFIV2-Val ( Integration Vector-tRNA -specific), using the sites and the site-specific integrase (FN3523_1033) of FhaGI-1, a chloramphenicol resistance cassette and a gene for counter selection of transformants against the vector backbone. We inserted the respective sites and genes into vector pUC57-Kana to allow for propagation in . The constructs generated a circular episomal form in which could be used to transform . where FIV-Val stably integrated site specifically into the tRNA gene of the genome, whereas pUC57-Kana is lost due to counter selection. Functionality of the new vector was demonstrated by the successfully complementation of a mutant strain. The vectors were stable and during host-cell infection without selective pressure. Thus, the vectors can be applied as a further genetic tool in research, expanding the present genetic tools by an integrative element. This new element is suitable to perform long-term experiments with different species.
ISSN:2235-2988
2235-2988
DOI:10.3389/fcimb.2018.00075