Earticle

초록

영어

Two piglets and one juvenile pig were used to investigate closely what types of cells express green fluorescent protein (GFP) and if any, whether the GFP-tagged cells could be used for stem cell transplantation research as a middle-sized animal model in bone marrow cells of recloned GFP pigs. Bone marrow cells were recovered from the tibia, and further analyzed with various cell lineage markers to determine which cell lineage is concurrently expressing visible GFP in each individual animal. In the three animals, visible GFP were observed only in proportions of the plated cells immediately after collection, showing 41, 2 and 91% of bone marrow cells in clones #1, 2 and 3, respectively. The intensity of the visible GFP expression was variable even in an individual clone depending on cell sizes and types. The overall intensities of GFP expression were also different among the individual clones from very weak, weak to strong. Upon culture for 14 days in vitro (14DIV), some cell types showed intensive GFP expression throughout the cells; in particular, in cytoskeletons and the nucleus, on the other hand. Others are shown to be diffused GFP expression patterns only in the cytoplasm. Finally, characterization of stem cell lineage markers was carried out only in the clone #3 who showed intensive GFP expression. SSEA-1, SSEA-3, CD34, nestin and GFAP were expressed in proportions of the GFP expressing cells, but not all of them, suggesting that GFP expression occur in various cell lineages. These results indicate that targeted insertion of GFP gene should be pursued as in mouse approach to be useful for stem cell research. Furthermore, cell- or tissue-specific promoter should also be used if GFP pig is going to be meaningful for a model for stem cell transplantation.

목차

ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
  Animals
  Recovery of Bone Marrow Cells
  Cell Culture
  Determination of GFP Expression in BMCs
  Immunocytochemistry for Cell Lineages
  Reverse Transcription-Polymerase Chain Reaction (RTPCR)and PCR of Genomic DNA
  Western Blotting Analysis for GFP
 RESULTS
  Different Extent of GFP Expression among the ReclonedGFP Pigs
  Cellular Distribution of GFP Expressed in GFP-positiveCells
  Expressions of Cell Surface Markers, SSEA-1, SSEA-3and CD34 in GFP-positive Cells
  Expressions of Neural Cell Lineage Markers in GFPpositiveCells
  Verification of Insertion and Expression of GFP Genein the Tissues of Recloned Pigs
 DISCUSSION
 REFERENCES

키워드

저자

• Kwang-Wook Park [ Department of Animal Science and Technology, Sunchon National University, Suncheon 540-742 and MGEN, Inc., #1101, World Meridian Venture Center, Seoul 153-781, Korea ]
• Sung Sik Choi [ Division of Life Sciences, Korea University, Seoul 136-701, Korea ]
• Dong Ho Lee [ Division of Life Sciences, Korea University, Seoul 136-701, Korea ]
• Hwang Lee [ Division of Life Sciences, Korea University, Seoul 136-701, Korea ]
• Seung Kyu Choi [ Division of Life Sciences, Korea University, Seoul 136-701, Korea ]
• Chang Sik Park [ Division of Animal Science & Resources, Research Center for Transgenic Cloned Pigs, Chungnam National University, Daejeon 305-764, Korea ]
• Sang Ho Lee [ Division of Life Sciences, Korea University, Seoul 136-701, Korea ] Corresponding author

참고문헌

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간행물

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• 간행물명

  Reproductive & developmental biology

• 간기

  계간

• pISSN

  1738-2432

• eISSN

  2288-0151

• 수록기간

  1977~2018

• 십진분류

  KDC 527 DDC 636