Journal of Embryo Transfer (한국수정란이식학회지)

The Korean Society of Embryo Transfer (한국수정란이식학회)
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DNA Methylation Change of H19 Differentially Methylated Region (DMR) in Day 35 of Cloned Pig Fetuses

돼지 체세포복제 35일령 태아에서 H19 메틸화 가변 영역의 DNA 메틸화 변화

  • Ko, Yeoung-Gyu (Animal Genetic Resources Station, National Institute of Animal Science, RDA) ;
  • Im, Gi-Sun (Animal Biotechnology Division, National Institute of Animal Science, RDA) ;
  • Hwang, Seong-Soo (Animal Biotechnology Division, National Institute of Animal Science, RDA) ;
  • Oh, Keon-Bong (Animal Biotechnology Division, National Institute of Animal Science, RDA) ;
  • Woo, Jae-Seok (Animal Biotechnology Division, National Institute of Animal Science, RDA) ;
  • Cho, Sang-Rae (Animal Genetic Resources Station, National Institute of Animal Science, RDA) ;
  • Choi, Sun-Ho (Animal Genetic Resources Station, National Institute of Animal Science, RDA) ;
  • Lee, Poong-Yeon (Animal Genetic Resources Station, National Institute of Animal Science, RDA) ;
  • Yeon, Sung-Heum (Animal Genetic Resources Station, National Institute of Animal Science, RDA) ;
  • Cho, Jae-Hyeon (Institute of Life Science, College of Veterinary Medicine, Gyeongsang National University)
  • 고응규 (농촌진흥청 국립축산과학원 가축유전자원시험장) ;
  • 임기순 (농촌진흥청 국립축산과학원 동물바이오공학과) ;
  • 황성수 (농촌진흥청 국립축산과학원 동물바이오공학과) ;
  • 오건봉 (농촌진흥청 국립축산과학원 동물바이오공학과) ;
  • 우제석 (농촌진흥청 국립축산과학원 동물바이오공학과) ;
  • 조상래 (농촌진흥청 국립축산과학원 가축유전자원시험장) ;
  • 최선호 (농촌진흥청 국립축산과학원 가축유전자원시험장) ;
  • 이풍연 (농촌진흥청 국립축산과학원 가축유전자원시험장) ;
  • 연성흠 (농촌진흥청 국립축산과학원 가축유전자원시험장) ;
  • 조재현 (경상대학교 수의과대학)
  • Received : 2011.02.26
  • Accepted : 2011.03.03
  • Published : 2011.03.31
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Abstract

This study was performed to identify the differentially methylated region (DMR) and to examine the mRNA expression of the imprinted H19 gene in day 35 of SCNT pig fetuses. The fetus and placenta at day 35 of gestation fetuses after natural mating (Control) or of cloned pig by somatic cell nuclear transfer (SCNT) were isolated from a uterus. To investigate the mRNA expression and methylation patterns of H19 gene, tissues from fetal liver and placenta including endometrial and extraembryonic tissues were collected. The mRNA expression was evaluated by real-time PCR and methylation pattern was analyzed by bisulfite sequencing method. Bisulfite analyses demonstrated that the differentially methylated region (DMR) was located between -1694 bp to -1338 bp upstream from translation start site of the H19 gene. H19 DMR (-1694 bp to -1338 bp) exhibits a normal mono allelic methylation pattern, and heavily methylated in sperm, but not in oocyte. In contrast to these finding, the analysis of the endometrium and/or extraembryonic tissues from SCNT embryos revealed a complex methylation pattern. The DNA methylation status of DMR Region In porcine H19 gene upstream was hypo methylated in SCNT tissues but hypermethylated in control tissues. Furthermore, the mRNA expression of H19 gene in liver, endometrium, and extraembryonic tissues was significantly higher in SCNT than those of control (p<0.05). These results suggest that the aberrant mRNA expression and the abnormal methylation pattern of imprinted H19 gene might be closely related to the inadequate fetal development of a cloned fetus, contributing to the low efficiency of genomic reprogramming.

Keywords

References

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