Journal of Animal Science and Technology

Korean Society of Animal Sciences and Technology (한국축산학회)
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Comparison of Vitrification and Slow Freezing for the Cryopreservation of Chicken Primordial Germ Cell (Ogye)

한국재래닭 (오계) 원시생식세포의 완만동결과 급속동결의 비교

  • Kim, Sung Woo (Animal Genetic Resources Station, National Institute of Animal Science, RDA) ;
  • Ko, Yeoung-Gyu (Animal Genetic Resources Station, National Institute of Animal Science, RDA) ;
  • Byun, Mijeong (Animal Genetic Resources Station, National Institute of Animal Science, RDA) ;
  • Do, Yoon Jung (Animal Genetic Resources Station, National Institute of Animal Science, RDA) ;
  • Han, Jae Yong (WCU Biomodulation Major, Department of Agricultural Biotechnology, Seoul National University) ;
  • Kim, Dong Hun (Animal Genetic Resources Station, National Institute of Animal Science, RDA) ;
  • Seong, Hwan-Hoo (Animal Genetic Resources Station, National Institute of Animal Science, RDA) ;
  • Kim, Hyun (Animal Genetic Resources Station, National Institute of Animal Science, RDA)
  • 김성우 (농촌진흥청 국립축산과학원 가축유전자원시험장) ;
  • 고응규 (농촌진흥청 국립축산과학원 가축유전자원시험장) ;
  • 변미정 (농촌진흥청 국립축산과학원 가축유전자원시험장) ;
  • 도윤정 (농촌진흥청 국립축산과학원 가축유전자원시험장) ;
  • 한재용 (서울대학교 동물자원과학과) ;
  • 김동훈 (농촌진흥청 국립축산과학원 가축유전자원시험장) ;
  • 성환후 (농촌진흥청 국립축산과학원 가축유전자원시험장) ;
  • 김현 (농촌진흥청 국립축산과학원 가축유전자원시험장)
  • Received : 2013.06.17
  • Accepted : 2013.09.13
  • Published : 2013.10.31
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Abstract

We sought to provide a method for freezing and preserving primordial germ cells, or an avian germ cell of a bird, as a material for developmental engineering or species preservation. The aim of this study was to compare the efficacy of slow freezing with a vitrification method for the cryopreservation of chicken primordial germ cells (PGCs). PGCs obtained from the germinal gonad of day 5.5-6 day (stage 28) cultured chick embryos, using the MACS method, were classified into two groups: slow freezing and vitrification. We examined the viability of PGCs after Cryopreservation. Four freezing methods were compared with each other, including the following: Method 1: The PGCs were frozen by a programmed freezer in a plastic straw, including 2.0 M ethylene glycol (EG) as cryoprotective additive (slow freezing) Method 2: The PGCs were vitrified in a plastic straw, including 8.0 M EG, plus 7% polyvinylpyrrolidone (PVP) (rapid freezing). Method 3: The slow freezing was induced with a cryotube including 2.0 M EG Method 4: The PGCs were frozen in a cryotube including 10% dimethyl suloxide (DMSO) (rapid freezing). After freezing and thawing, survival rates of the frozen-thawed PGCs from Method 1 to 4were 76.4%, 70.6%, 80.5% and 78.1% (p<0.05), respectively. The slow freezing ($-80^{\circ}C$ programmed freezer) method may provide better survival rates of frozen-thawed PGCs than the vitrification method for the cryopreservation of PGCs. Therefore, these systems may contribute to the cryopreservation of a rare avian species.

동결 닭 PGCs의 생식계열 키메라를 이용한 생체에의 복원을 실용화 하기 위해서는, 닭 PGCs의 동결보존기술의 향상에 의해 동결 및 융해 후의 많은 생존세포를 확보 하는 것과, 생식계열 키메라의 제작효율을 높이는 것이 반드시 필요하다. 닭 PGCs는 배양 5.5일령의 닭 원시생식선으로부터 채취하고, ACS 방법에 의해서 순수 닭 PGCs를 분리했다. 닭 PGCs의 동결보존실험결과 다음의 4종류의 동결방법을 각각 비교 검토했다. 1. 플라스틱 스트로에 의한 완만동결법 (SF), 동결보호물질은 2M 에틸렌 글리콜 (EG), 2. 스트로에 의한 급속동결법 (RF), 8M EG + 7% PVP, 3. 동결용 Cryotube에 의한 SF, 2M EG, 4. 튜브에 의한 SF, 10% DMSO. 동결 및 융해 후의 PGCs의 생존율은 각각 76.4%, 70.6%, 80.5%, 78.1%로 관찰되었다.

Keywords

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