Abstract
BackgroundDNA methylation of cytosine residues in CpG dinucleotide controls gene expression and dramatically changes during development. Its pattern is disrupted in cloned animals suggesting incomplete reprogramming during somatic cell nuclear transfer (the first reprogramming). However, the second reprogramming occurs in the germ cells and epigenetic errors in somatic cells of cloned animals should be erased. To analyze the DNA methylation changes on the spermatogenesis of bulls, we measured DNA methylation levels of three repetitive elements in blastocysts, blood and sperm.MethodsDNA from PBLs (peripheral blood leukocytes), sperm and individual IVF (in vitro fertilized) and parthenogenetic blastocysts was isolated and bisulfite converted. Three repetitive elements; Satellite I, Satellite II and art2 sequences were amplified by PCR with specific pairs of primers. The PCR product was then cut by restriction enzymes and analyzed by agarose gel electrophoresis for determining the DNA methylation levels.ResultsBoth Satellite I and Satellite II sequences were highly methylated in PBLs, whereas hypo-methylated in sperm and blastocysts. The art2 sequence was half methylated both in PBLs and sperm but less methylated in blastocysts. There was no difference in DNA methylation levels between IVF and parthenogenetic blastocysts.ConclusionsThese results suggest that there is a dynamic change of DNA methylation during embryonic development and spermatogenesis in cattle. Satellite I and Satellite II regions are methylated during embryogenesis and then de-methylated during spermatogenesis. However, art2 sequences are not de-methylated during spermatogenesis, suggesting that this region is not reprogrammed during germ cell development. These results show dynamic changes of DNA methylation levels during bovine embryogenesis, especially genome-wide reprogramming in germ cells.
Highlights
DNA methylation of cytosine residues in CpG dinucleotide controls gene expression and dramatically changes during development
Art2 sequences are not de-methylated during spermatogenesis, suggesting that this region is not reprogrammed during germ cell development
These results show dynamic changes of DNA methylation levels during bovine embryogenesis, especially genome-wide reprogramming in germ cells
Summary
DNA methylation of cytosine residues in CpG dinucleotide controls gene expression and dramatically changes during development. Cytosine residues in CpG dinucleotide pairs are selectively methylated by DNA methyltransferases and these methylation patterns are maintained throughout cell division. DNA methylation alters gene expression patterns in cells and is crucial for normal mammalian development [1,2]. Repetitive elements such as centromeric repeats and methylation patterns were observed in many kinds of tumour cells [4]. We report the dynamic changes of DNA methylation patterns at three repetitive sequences in bovine blastocysts, somatic cells and sperm
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