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Epigenetic and Genomic Imprinting Analysis in Nuclear Transfer Derived Bos gaurus/Bos taurus Hybrid Fetuses¹

Department of Veterinary Anatomy and Public Health,⁴ College of Veterinary Medicine, Texas A&M University, College Station, Texas 77840 Department of Population Health and Pathobiology⁵ Department of Molecular Biomedical Science,⁶ College of Veterinary Medicine, North Carolina State University, Raleigh, North Carolina 27606 Department of Animal Science,⁷ College of Agriculture and Life Sciences, North Carolina State University, Raleigh, North Carolina 27695-7621 Viagen Inc.,⁸ College Station, Texas 77843

Somatic cell nuclear transfer (NT) in cattle is an inefficient process, whereby the production of calves is hindered by low pregnancy rates as well as fetal and placental abnormalities. Interspecies models have been previously used to facilitate the identification of single nucleotide polymorphisms (SNPs) within coding regions of genes to discriminate between parental alleles in the offspring. Here we report the use of a bovine interspecies model (Bos gaurus x Bos taurus) for the assessment and characterization of epigenetic modifications and genomic imprinting in Day 40-old female NT-derived fetuses and placenta. Analysis of NT and control pregnancies indicated disruption of genomic imprinting at the X inactivation-specific transcript (XIST) locus in the chorion, but not the fetus of clones, whereas proper allelic expression of the insulin-like growth factor II (IGF2) and gene trap locus 2 (GTL2) loci was maintained in both the fetus and placenta. Analysis of the XIST differentially methylated region (DMR) in clones indicated normal patterns of methylation; however, bisulfite sequencing of the satellite I repeat element and epidermal cytokeratin promoter indicated hypermethylation in the chorion of clones when compared with controls. No differences were detected in methylation levels in the fetus proper. These results indicate that the nuclear transfer process affects gene expression patterns in the trophectoderm- and inner cell mass-derived tissues to different extents.

¹ Supported by NIH grant HL51587 and a Texas A&M University, College of Veterinary Medicine Signature grant.

² Correspondence: Jorge A. Piedrahita, Department of Molecular Biomedical Science, College of Veterinary Medicine, North Carolina State University, 611 Hutton St., Raleigh, NC 27606; FAX: 919 515 4237; jorge_piedrahita{at}ncsu.edu

³ Current address: Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX 77030