Analysis of mouse monozygotic twin blastocysts produced in two different culture media

Following fertilization in mammals, it is generally accepted that totipotent cells are exclusive to the zygote and to each of the two blastomeres originating from the first mitotic division. We counter that this classic view needs to be revised, because we have presented compelling evidence the sister blastomeres are both totipotent in only a subset of 2-cell stage mouse embryos (PMID 28811525). Building on our previous findings, we here ask the question if the differences between sister blastomeres can be modulated experimentally. To this end, we separate the sister blastomeres, yielding monozygotic twins. We culture these twins in two different media (GM501 vs. Sage 1 step), to see if the mRNA ratios of twin blastocysts lie more far apart from each other when culture took place in different media as compared to culture in the same medium (GSE90674). Transcriptome analysis followed by intra-pair mRNA ratio analysis revealed differences between the blastocysts of the same monozygotic pair. Some of the differences were sensitive to the choice of culture conditions, while other differences were insensitive. After in vivo zygote production and short (2 hours) culture in KSOM(aa) medium, 2-cell embryos were split, and the individual blastomeres were cultured in parallel in GM501 vs. Sage 1 step medium. Resultant monozygotic twins were sampled at the blastocyst stage, respecting the original pair associations (e.g. blastocyst '1a' and blastocyst '1b' both derived from same 2-cell embryo number '1'). Samples were subjected to Affymetrix microarray analysis, performed by Anika Witten c/o Institut fuer Humangenetik, Universitaetsklinikum Muenster.