Abstract
Somatic cell nuclear transfer (SCNT), also known as somatic cell cloning, is a commonly used technique to study epigenetic reprogramming. Although SCNT has the advantages of being safe and able to obtain pluripotent cells, early developmental arrest happens in most SCNT embryos. Overcoming epigenetic barriers is currently the primary strategy for improving reprogramming efficiency and improving developmental rate in SCNT embryos. In this study, we analyzed DNA methylation profiles of in vivo fertilized embryos and SCNT embryos with different developmental fates. Overall DNA methylation level was higher in SCNT embryos during global de-methylation process compared to in vivo fertilized embryos. In addition, promoter region, first intron and 3′UTR were found to be the major genomic regions that were hyper-methylated in SCNT embryos. Surprisingly, we found the length of re-methylated region was directly related to the change of methylation level. Furthermore, a number of genes including Dppa2 and Dppa4 which are important for early zygotic genome activation (ZGA) were not properly activated in SCNT embryos. This study comprehensively analyzed genome-wide DNA methylation patterns in SCNT embryos and provided candidate target genes for improving efficiency of genomic reprogramming in SCNT embryos. Since SCNT technology has been widely used in agricultural and pastoral production, protection of endangered animals, and therapeutic cloning, the findings of this study have significant importance for all these fields.
Highlights
Somatic cell nuclear transfer (SCNT) technology can restore pluripotency of mature somatic cells, and generate new offspring by transplanting somatic cell nuclei into enucleated oocytes (Rodriguez-Osorio et al, 2012; Zuo et al, 2017)
In order to have a deeper insight into the relationship between DNA methylation and low developmental potential of SCNT embryos, we conducted genome-wide DNA methylation analysis based on whole-genome bisulfite sequencing (WGBS) data obtained from different developmental stages of SCNT embryos, including: zygote, 2-cell arrest (NA2), 2-cell to blastocyst (NB2), 4-cell arrest (NA4), 4-cell to blastocyst (NB4); and in vivo fertilized embryos including: in vivo fertilized 2-cell (WT2), in vivo fertilized 4-cell (WT4); and Cumulus cell (CC) (Gao et al, 2018)
We found that overall DNA methylation level was higher in SCNT embryos during global de-methylation process compared to in vivo fertilized embryos
Summary
Somatic cell nuclear transfer (SCNT) technology can restore pluripotency of mature somatic cells, and generate new offspring by transplanting somatic cell nuclei into enucleated oocytes (Rodriguez-Osorio et al, 2012; Zuo et al, 2017). Gurdon transplanted Xenopus laevis epithelial cells in tadpole stage into eggs with damaged nuclei. A small fraction of the eggs developed into normal embryos, demonstrating that differentiated somatic cells can still restore pluripotency (Gurdon, 1962). Accumulating evidences suggested that incomplete reprogramming of epigenetic status including chromatin accessibility, histone modifications and DNA methylation of the donor cell genome, is a determining factor for the low cloning efficiency (Cantone and Fisher, 2013; Niemann, 2016)
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