Improvement of a porcine somatic cell nuclear transfer technique by optimizing donor cell and recipient oocyte preparations

Gabsang Lee, Sang Hwan Hyun, Hye Soo Kim, Dae Young Kim, So Hyun Lee, Jeong Mook Lim, Eun Song Lee, Sung Keun Kang, Byeong Chun Lee, Woo Suk Hwang

Research output: Contribution to journalArticle

Abstract

This study was conducted to improve a porcine somatic cell nuclear transfer (SCNT) technique by optimizing donor cell and recipient oocyte preparations. Adult and fetal fibroblasts, and cumulus and oviduct cells were used as donor cells, and in vivo- and in vitro-matured oocytes were employed as recipient oocytes. The percentages of fusion and development to the blastocyst stage, the ratio of blastocysts to 2-cell embryos, and cell number of blastocysts were monitored as experimental parameters. In Experiment 1, donor cells of four different types were transferred to enucleated oocytes matured in vitro, and more (P < 0.05) blastocysts were derived from SCNT of fetal fibroblasts than from that of other cells (15.9% versus 3.1-7.9%). For SCNT using fetal fibroblasts, increasing the number of subcultures up to 15 times did not improve developmental competence to the blastocyst stage (12.2-16.7%). In Experiment 2, fetal fibroblasts were transferred to enucleated oocytes that matured in vivo or in vitro. When parthenogenetic activation of both types of oocytes was conducted as a preliminary control treatment, a signifcant increase in blastocyst formation was found for in vivo-matured compared with in vitro-matured oocytes (36.4% versus 29.5%). However, no improvement was achieved in SCNT using in vivo-matured oocytes. In conclusion, the type of donor somatic cell is important for improving development after porcine SCNT, and fetal fibroblasts were the most effective among examined cells. A system with good reproducibility has been established using fetal fibroblasts as the donor karyoplast after subculturing 1-10 times, and using both in vivo and in vitro-matured oocytes as the recipient cytoplast.

Original languageEnglish (US)
Pages (from-to)1949-1957
Number of pages9
JournalTheriogenology
Volume59
Issue number9
DOIs
StatePublished - Jan 1 2003
Externally publishedYes

Fingerprint

Nuclear Transfer Techniques
nuclear transplantation
Oocytes
oocytes
Swine
blastocyst
swine
fibroblasts
Blastocyst
somatic cells
Fibroblasts
cells
Cumulus Cells
oviducts
Oviducts
reproducibility
embryo (animal)
Mental Competency

Keywords

  • Karyoplast type
  • Oocyte source
  • Parthenogenetic activation
  • Porcine
  • Reconstructed embryos

ASJC Scopus subject areas

  • Small Animals
  • Food Animals
  • Animal Science and Zoology
  • Equine

Cite this

Improvement of a porcine somatic cell nuclear transfer technique by optimizing donor cell and recipient oocyte preparations. / Lee, Gabsang; Hyun, Sang Hwan; Kim, Hye Soo; Kim, Dae Young; Lee, So Hyun; Lim, Jeong Mook; Lee, Eun Song; Kang, Sung Keun; Lee, Byeong Chun; Hwang, Woo Suk.

In: Theriogenology, Vol. 59, No. 9, 01.01.2003, p. 1949-1957.

Research output: Contribution to journalArticle

Lee, G, Hyun, SH, Kim, HS, Kim, DY, Lee, SH, Lim, JM, Lee, ES, Kang, SK, Lee, BC & Hwang, WS 2003, 'Improvement of a porcine somatic cell nuclear transfer technique by optimizing donor cell and recipient oocyte preparations', Theriogenology, vol. 59, no. 9, pp. 1949-1957. https://doi.org/10.1016/S0093-691X(02)01294-3
Lee, Gabsang ; Hyun, Sang Hwan ; Kim, Hye Soo ; Kim, Dae Young ; Lee, So Hyun ; Lim, Jeong Mook ; Lee, Eun Song ; Kang, Sung Keun ; Lee, Byeong Chun ; Hwang, Woo Suk. / Improvement of a porcine somatic cell nuclear transfer technique by optimizing donor cell and recipient oocyte preparations. In: Theriogenology. 2003 ; Vol. 59, No. 9. pp. 1949-1957.
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abstract = "This study was conducted to improve a porcine somatic cell nuclear transfer (SCNT) technique by optimizing donor cell and recipient oocyte preparations. Adult and fetal fibroblasts, and cumulus and oviduct cells were used as donor cells, and in vivo- and in vitro-matured oocytes were employed as recipient oocytes. The percentages of fusion and development to the blastocyst stage, the ratio of blastocysts to 2-cell embryos, and cell number of blastocysts were monitored as experimental parameters. In Experiment 1, donor cells of four different types were transferred to enucleated oocytes matured in vitro, and more (P < 0.05) blastocysts were derived from SCNT of fetal fibroblasts than from that of other cells (15.9{\%} versus 3.1-7.9{\%}). For SCNT using fetal fibroblasts, increasing the number of subcultures up to 15 times did not improve developmental competence to the blastocyst stage (12.2-16.7{\%}). In Experiment 2, fetal fibroblasts were transferred to enucleated oocytes that matured in vivo or in vitro. When parthenogenetic activation of both types of oocytes was conducted as a preliminary control treatment, a signifcant increase in blastocyst formation was found for in vivo-matured compared with in vitro-matured oocytes (36.4{\%} versus 29.5{\%}). However, no improvement was achieved in SCNT using in vivo-matured oocytes. In conclusion, the type of donor somatic cell is important for improving development after porcine SCNT, and fetal fibroblasts were the most effective among examined cells. A system with good reproducibility has been established using fetal fibroblasts as the donor karyoplast after subculturing 1-10 times, and using both in vivo and in vitro-matured oocytes as the recipient cytoplast.",
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