Title page for ETD etd-07312006-124855


Type of Document Dissertation
Author Moisan, Allison E.
Author's Email Address amoisa1@lsu.edu
URN etd-07312006-124855
Title Preservation of Gametes Using Vitrification and Dehydration
Degree Doctor of Philosophy (Ph.D.)
Department Animal Science (Animal, Dairy, & Poultry Sciences)
Advisory Committee
Advisor Name Title
Robert Godke Committee Chair
John Lynn Committee Member
Kennith Bondioli Committee Member
Stanley Leibo Committee Member
Jill Johnson Dean's Representative
Keywords
  • feline cryopreservation
  • bovine cryopreservation
  • sperm dehydration
  • oocyte vitrification
  • freeze-dried sperm
Date of Defense 2006-03-20
Availability unrestricted
Abstract
Of the 36 species of felines in the world, all except the domestic cat are listed as endangered or threatened. To preserve the genetic diversity of felines and other species, genome resource banks have been established. Due to limited availability of germ cells for research, studies must use models to optimize the techniques before they are applied to endangered species. In this study, preservation of oocytes and spermatozoa was examined using the bovine as a model for felines. In the first series of experiments, bovine and feline oocytes were dehydrated, vitrified, warmed and cultured to assess their ability to undergo embryonic development using a choline-based medium (CJ2) for vitrification and warming solutions preparation as well as the standard sodium based media. In the second series of experiments, feline spermatozoa were dehydrated using air- and freeze-drying as alternative methods to standard cryopreservation. Assessment was done by examining embryonic development after intracytoplasmic sperm injection (ICSI) and DNA integrity of the dehydrated spermatozoa using the comet assay. In the second series of experiments, bovine and feline oocytes behaved osmotically in response to increasingly concentrated solutions. However, vitrified-warmed bovine oocytes had significantly higher cleavage and blastocyst rates compared with their feline counterparts and development using CJ2 medium was similar to the standard media used for cattle but was detrimental to feline oocytes. In the third experiment, cleavage and blastocyst development of feline oocytes injected with cat spermatozoa preserved using air- and freeze-drying was observed. Also, exposure to the dehydration solution and vitrification did not induce DNA damage but the process of freeze-drying did have significantly higher levels compared with controls. Air-dried sperm did not decondense. In conclusion, the use of bovine oocytes as a model for feline oocytes was successful. Both bovine and feline oocytes responded similarly to dehydration and vitrification, except when processed using CJ2 medium. Furthermore, feline spermatozoa can be preserved using dehydration as demonstrated by their ability to produce blastocysts. This study has encouraging results for germ cell preservation. However, the efficiency of these procedures must be improved before they can be used as alternative methods of preservation in endangered species.
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