Effects of Trichostatin A on In Vitro Development and DNA Methylation Level of the Satellite I Region of Swamp Buffalo (Bubalus bubalis) Cloned Embryos

Trichostatin A (TSA), a histone deacetylase inhibitor, has been widely used to improve the cloning efficiency in severalspecies. This brings our attention to investigation of the effects of TSA on developmental potential of swamp buffalo clonedembryos. Swamp buffalo cloned embryos were produced by electrical pulse fusion of male swamp buffalo fibroblasts with swampbuffalo enucleated oocytes. After fusion, reconstructed oocytes were treated with 0, 25 or 50 nM TSA for 10 h. The resultsshowed that there was no significant difference in the rates of fusion (82–85%), cleavage (79–84%) and development to the8-cell stage (59–65%) among treatment groups. The highest developmental rates to the morula and blastocyst stages of embryoswere found in the 25 nM TSA-treated group (42.7 and 30.1%, respectively). We also analyzed the DNA methylation level in thesatellite I region of donor cells and in in vitro fertilized (IVF) and cloned embryos using the bisulfiteDNA sequencing method. The results indicated that the DNA methylation levels in cloned embryos were significantly higher thanthose of IVF embryos but approximately similar to those of donor cells. Moreover, there was no significant difference in themethylation level among TSA-treated and untreated cloned embryos. Thus, TSA treatments at 25 nM for 10 h could enhance thein vitro developmental potential of swamp buffalo cloned embryos, but no beneficial effect on the DNAmethylation level was observed.