Knockout of Babesia bovis rad51 ortholog and its complementation by expression from the BbACc3 artificial chromosome platform

PloS One
Erin A. MackDavid R Allred

Abstract

Babesia bovis establishes persistent infections of long duration in cattle, despite the development of effective anti-disease immunity. One mechanism used by the parasite to achieve persistence is rapid antigenic variation of the VESA1 cytoadhesion ligand through segmental gene conversion (SGC), a phenomenon thought to be a form of homologous recombination (HR). To begin investigation of the enzymatic basis for SGC we initially identified and knocked out the Bbrad51 gene encoding the B. bovis Rad51 ortholog. BbRad51 was found to be non-essential for in vitro growth of asexual-stage parasites. However, its loss resulted in hypersensitivity to methylmethane sulfonate (MMS) and an apparent defect in HR. This defect rendered attempts to complement the knockout phenotype by reinsertion of the Bbrad51 gene into the genome unsuccessful. To circumvent this difficulty, we constructed an artificial chromosome, BbACc3, into which the complete Bbrad51 locus was inserted, for expression of BbRad51 under regulation by autologous elements. Maintenance of BbACc3 makes use of centromeric sequences from chromosome 3 and telomeric ends from chromosome 1 of the B. bovis C9.1 line. A selection cassette employing human dihydrofolate reductase enable...Continue Reading

References

Jul 1, 1992·Molecular and Biochemical Parasitology·S M Kamper, A F Barbet
Jun 11, 1992·Nucleic Acids Research·M J van den HoffW H Lamers
Oct 5, 1990·Journal of Molecular Biology·S F AltschulD J Lipman
Dec 1, 1989·Proceedings of the National Academy of Sciences of the United States of America·C RothH Eisen
Jan 1, 1984·Annual Review of Biochemistry·E H Blackburn
Jul 1, 1993·Molecular and Biochemical Parasitology·D R AllredK P Ahrens
Feb 15, 1996·Nucleic Acids Research·P K PatnaikG A Cross
Sep 1, 1997·Nucleic Acids Research·S F AltschulD J Lipman
Jul 17, 1998·The New England Journal of Medicine·P J KrauseD H Persing
Feb 5, 2000·The Journal of Immunology : Official Journal of the American Association of Immunologists·R M O'Connor, D R Allred
Dec 20, 2000·Bioinformatics·K RutherfordB Barrell
Feb 14, 2002·International Journal for Parasitology·Manoj T DuraisinghAlan F Cowman
Mar 30, 2002·The Journal of Biological Chemistry·Jeremy M StarkMaria Jasin
Oct 18, 2003·Current Opinion in Hematology·Ritchard G Cable, David A Leiby
Jan 30, 2004·Molecular and Biochemical Parasitology·David R Allred, Basima Al-Khedery
Jun 25, 2004·Nucleic Acids Research·Aron Marchler-Bauer, Stephen H Bryant
Jun 25, 2004·Nucleic Acids Research·David E KimDavid Baker
Jul 6, 2004·Nature Structural & Molecular Biology·Adam B ConwayPhoebe A Rice
Jul 21, 2004·Journal of Computational Chemistry·Eric F PettersenThomas E Ferrin
Sep 24, 2004·International Journal for Parasitology·Carlos E SuarezTerry F McElwain
Dec 6, 2005·Nucleic Acids Research·Chris Proudfoot, Richard McCulloch
Mar 7, 2006·Veterinary Parasitology·David R Allred, Basima Al-Khedery
Mar 15, 2006·Veterinary Parasitology·Erik de VriesMatthew Berriman
May 9, 2006·International Journal for Parasitology·Carlos E SuarezTerry F McElwain
May 27, 2006·International Journal for Parasitology·Patricia A ConradBarbara L Herwaldt
Apr 20, 2007·Experimental Parasitology·Audrey O T LauTerry F McElwain
Oct 24, 2007·PLoS Pathogens·Kelly A BraytonVishvanath M Nene
Jan 10, 2008·Clinical Infectious Diseases : an Official Publication of the Infectious Diseases Society of America·Peter J KrauseAndrew Spielman
Oct 4, 2008·International Journal for Parasitology·Carlos E Suarez, Terry F McElwain
Apr 14, 2009·Nature Methods·Daniel G GibsonHamilton O Smith
Jul 7, 2009·Mutation Research·Takeo OhnishiAkihisa Takahashi
Aug 25, 2009·Proteins·Srivatsan RamanDavid Baker

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Methods Mentioned

BETA
transfection
PCR
light microscopy

Software Mentioned

Artemis
Robetta
Bio Main Workbench
Chimera
CLC
Psi
Chimera Matchmaker
Delta
BLAST

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