Isolation, culture and characterisation of somatic cells derived from semen and milk of endangered sheep and eland antelope
L. Nel-Themaat A B , M. C. Gómez A B , P. Damiani B , G. Wirtu B C , B. L. Dresser B D , K. R. Bondioli A F , L. A. Lyons E , C. E. Pope B and R. A. Godke A CA Department of Animal Sciences, Louisiana State University Agricultural Center, Baton Rouge, LA 70803, USA.
B Audubon Center for Research of Endangered Species, 14001 River Road, New Orleans, LA 70131, USA.
C Department of Veterinary Clinical Sciences, Louisiana State University School of Veterinary Medicine, Baton Rouge, LA 70803, USA.
D Department of Biological Sciences, University of New Orleans, New Orleans, LA 70148, USA.
E School of Veterinary Medicine, University of California—Davis, Davis, CA 95616, USA.
F Corresponding author. Email: Kbondioli@agcenter.lsu.edu
Reproduction, Fertility and Development 19(4) 576-584 https://doi.org/10.1071/RD06153
Submitted: 8 November 2006 Accepted: 19 March 2007 Published: 7 May 2007
Abstract
Semen and milk are potential sources of somatic cells for genome banks. In the present study, we cultured and characterised cells from: (1) cooled sheep milk; (2) fresh, cooled and frozen–thawed semen from Gulf Coast native (GCN) sheep (Ovis aries); and (3) fresh eland (Taurotragus oryx) semen. Cells attached to the culture surface from fresh (29%), cooled (43%) and slow-frozen (1°C/min; 14%) ram semen, whereas no attachment occurred in the fast-frozen (10°C/min) group. Proliferation occurred in fresh (50%) and cooled (100%) groups, but no cells proliferated after passage 1 (P1). Eland semen yielded cell lines (100%) that were cryopreserved at P1. In samples from GCN and cross-bred milk, cell attachment (83% and 95%, respectively) and proliferation (60% and 37%, respectively) were observed. Immunocytochemical detection of cytokeratin indicated an epithelial origin of semen-derived cells, whereas milk yielded either fibroblasts, epithelial or a mixture of cell types. Deoxyribonucleic acid microsatellite analysis using cattle-derived markers confirmed that eland cells were from the semen donor. Eland epithelial cells were transferred into eland oocytes and 12 (71%), six (35%) and two (12%) embryos cleaved and developed to morulae or blastocyst stages, respectively. In conclusion, we have developed a technique for obtaining somatic cells from semen. We have also demonstrated that semen-derived cells can serve as karyoplast donors for nuclear transfer.
Additional keywords: cell culture, cryopreservation, ejaculate, epithelial, Gulf Coast native sheep, nuclear transfer.
Acknowledgements
The authors extend their gratitude to Tray Harding and the LSU Agricultural Center Sheep Research Station team for providing the sheep and helping with sample collection. The authors also acknowledge Dr Alex Cole, Contessa Knight and the animal keepers from the Audubon Research Center Species Survival Center for helping with the eland semen collections.
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