Patterns /Expression Levels of Imprinted Genes in Clones

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Genetically imprinted genes are only expressed from either the maternal or paternal allele. This is due to differential "marks" established on the DNA of sperm and oocytes during gametogenesis, which are maintained throughout embryogenesis and subsequent development. In somatic cloning by nuclear transfer, however, no separate gametes with differentially marked genes are involved, both sets of chromosomes are derived from the same donor cell. Therefore, it is of interest to find out how the expression of imprinted genes is regulated in somatic clones. This question is intriguing because signs of imprinting disruption have been observed in cloned cattle regardless of donor cell type or donor age. Developmental defects such as high fetal and neonatal deaths and large calves are reminiscent of experimentally created imprinting disruptions (biallelic expression of imprinted genes) in mice and naturally occurring imprinting diseases in humans. Therefore, we hypothesize that some of the defects consistently observed in somatic clones are likely associated with aberrant expressions of imprinted genes after cloning. Our long-term objective is to understand whether and how imprinted genes are altered in clones. Our specific aims of this proposal are 1) to determine whether imprinting patterns are disrupted in cloned animals derived from adult somatic cells, and 2) to determine if the levels of mRNA for imprinted genes are altered in somatic clones. These specific aims are capitalized on our preliminary results that 1) we have collected samples from eleven live or dead somatic cattle clones which showed signs of imprinting disruptions, and 2) we have established polymorphic assays for IGF2 and IGF2R in bovine and our donor is heterozygous in both loci. Finding fromthese explorative studies will not only provide insights into the low efficiency of somatic cloning but also significantly further our understanding of the establishment and regulation of genetic imprinting during mammalian development.