Title page for ETD etd-09052007-124840

Type of Document Master's Thesis
Author DeRouen Polito, Amanda Paige
Author's Email Address aderou2@lsu.edu
URN etd-09052007-124840
Title Depletion of 32-Kbp Circular Plasmids from Borrelia burgdorferi
Degree Master of Science (M.S.)
Department Pathobiological Sciences (Veterinary Medical Sciences)
Advisory Committee
Advisor Name Title
Fang-Ting Liang Committee Chair
Kevin R. Macaluso Committee Member
Ronald L. Thune Committee Member
  • lyme disease
  • borrelia burgdorferi
  • cp32 plasmids
Date of Defense 2007-06-28
Availability unrestricted
The Lyme disease spirochete Borrelia burgdorferi has a very unusual genome composed of one linear chromosome and up to 21 linear and circular plasmids. Several plasmids are known to be important either for mammalian infection or tick colonization. A single spirochete harbors up to 7 different cp32 plasmids; however, nothing is known about their role in mammalian infection. The plasmids in this family are well maintained during in vitro cultivation, making it difficult to study their functions. To effectively deplete the plasmids, an 8kbp fragment containing essential elements for replication and partitioning in B. burgdorferi was amplified from one of the cp32 plasmids, cp32-3, and cloned into the vector pGE22 that carries a gentamycin resistance cassette and essential elements for replication in Escherichia coli. The resulting construct, pG22cp32-3plus, was electroporated into borrelial cells. By increasing gentamycin selection pressure, the spirochetes were forced to lose the corresponding cp32 plasmid. This strategy can be used to knock out other members of the cp32 family.
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