Improved technology of mouse somatic cell nuclear transfer that greatly increases cloning efficiency

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Production rate of cloned mice still is very low, which is thought to be attributed to abnormal epigenetic modifications and X-chromosomal inactivation (XCI) status in somatic cell nuclear transfer (SCNT) embryos. Here, we developed novel mouse SCNT technologies as follows; 1) treatment with valproi acid for 24 h from the 4-cell stage of SCNT embryos, which improved expression of Oct4 and trimethylated histone H3 lysine 27, a marker of the XCI status in SCNT blastocysts, 2) intracytoplasmic injection of deionized BSA after SCNT, which promoted expression of acetylated histone H3 lysine 9 and acetylated histone H4 lysine 12 at the pronuclear stage and production of cloned offspring, and 3) treatments with tricostatine A for 8 h from the beginning of activation of SCNT oocytes followed by vitamin C for 7 h, which promoted the oxidation of 5-methylcytocine to 5-hydroxymethylcytosine in pronuclear DNA of SCNT embryos and led to greatly increased production of cloned offspring.