Title

Development and subsequent reproductive performance of cloned cattle derived from adult somatic cells

Date of Completion

January 2003

Keywords

Biology, Genetics|Biology, Animal Physiology|Agriculture, Animal Culture and Nutrition

Degree

Ph.D.

Abstract

Development to blastocyst following nuclear transfer (NT) is dependent on the donor cell's ability to reprogram its genome to that of a zygote. This reprogramming step is inefficient and may be dependent on a number of factors, including chromatin organization. Donor cell type, cell cycle stage, and passage number of cultured cells all affect the developmental potential of cloned embryos through their chromatin status. Furthermore, successful somatic cell cloning has resulted in small numbers of live animals due to the low efficiency of the overall cloning process. We investigated the differences in histone acetylation of bovine fibroblast and cumulus cells at various passages and cell cycle stages. Our results show that stage of cell cycle, cell type, and numbers of cell passages all had an effect on histone content. In subsequent studies, Trichostatin A (TSA; 0–5 μM), a histone de-acetylase inhibitor, was used to increase histone acetylation in an attempt to improve donor cell re-programmability. Treatment of donor cells with 0.08 μM TSA increased blastocyst development compared to controls (35.1% vs. 25.1%). These results indicate that partial erasure of pre-existing epigenetic marks of donor cells improves subsequent development of cloned embryos. Since the in vivo development of cloned embryos remains relatively low, these observations raise the question of whether the genome of somatic clones is completely re-programmed and whether healthy clones from aged animals can grow and reproduce normally. By measuring the age at puberty, follicle growth characteristics and reproductive hormone profiles of cloned and control heifers; our results demonstrate that clones from an aged adult have normal reproductive development. ^