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BOR - Papers in Press, published online ahead of print May 23, 2007.
Biol Reprod 2007, 10.1095/biolreprod.106.058172
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BIOLOGY OF REPRODUCTION 77, 384–394 (2007)
DOI: 10.1095/biolreprod.106.058172
© 2007 by the Society for the Study of Reproduction, Inc.

Red Deer Cloned from Antler Stem Cells and Their Differentiated Progeny1

Debra K Berg 3, Chunyi Li 4, Geoff Asher 5, David N Wells 3, and Björn Oback 2 3

Reproductive Technologies,3 Ruakura Research Centre, AgResearch Ltd., Hamilton, New Zealand Growth and Development4 and Agricultural Systems,5 Invermay Agricultural Centre, AgResearch Ltd., Mosgiel, New Zealand

ABSTRACT

The significance of donor cell differentiation status for successful cloning by somatic cell nuclear transfer (SCNT) is unclear. Here, we cloned a new species, red deer (Cervus elaphus), from multipotent antler stem cells and their differentiated progeny. Cultured donor cell lines from male antlerogenic periosteum (AP) were left undifferentiated or chemically induced to initiate osteogenesis or adipogenesis. Based on their morphology and marker gene expression profile, donor cells were classified as undifferentiated AP cells, presumptive osteoblasts, or adipocytes. Adipocytes upregulated adipogenic markers procollagen type I alpha 2 (COL1A2), peroxisome proliferator-activated receptor gamma 2 (PPARG), and gylceraldehyde-3-phosphate dehydrogenase (GAPDH), and downregulated antlerogenic transcripts POU-domain class 5 transcription factor (POU5F1) and parathyroid hormone (PTH)-like hormone (PTHLH). Despite differences prior to NT, transcript abundance of donor-specific markers COL1A2, PPARG, GAPDH, and POU5F1 did not differ significantly in cloned blastocysts (P = 0.10, 0.50, 0.61, and 0.16, respectively). However, donor cell and blastocyst expression levels were completely different for most genes analyzed, indicating their successful reprogramming. The type of donor cell used for NT (AP, bone, and fat cells), had no effect on in vitro development to blastocysts (93 [38%] of 248 vs. 32 [44%] of 73 vs. 59 [32%] of 183, respectively). Likewise, development to weaning was not significantly different between the three cell types (2 [4%] of 46 vs. 2 [29%] of 7 vs. 4 [13%] of 31, for AP vs. bone vs. fat, respectively). Microsatellite DNA analysis confirmed that the eight cloned red deer calves were genetically identical to the cells used for NT.

assisted reproductive technology, developmental biology, early development, embryo, pregnancy

FOOTNOTES

1Supported by the New Zealand Foundation for Research, Science, and Technology.

Correspondence: 2FAX: 64 7 838 5536; e-mail: bjorn.oback{at}agresearch.co.nz






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Copyright © 2007 by the Society for the Study of Reproduction.