Project description:10.1021/acs.analchem.6b01921
Article: Single Cell Proteomics Using Frog (Xenopus laevis) Blastomeres Isolated from Early Stage Embryos, Which Form a Geometric Progression in Protein Content.
Project description:To determine blastomere fate and embryonic genome activation (EGA) at 5- to 8-cell stage human embryos by global gene expression profile of amplified cDNA from blastomeres at the single cell level Forty-nine blastomeres from 5-, 6- and 8-cell human embryos were analyzed through whole genome wide analysis following an efficient cDNA amplification protocol (Kurimoto et al., 2007) with slight modifications. Single biopsied blastomeres were also compared with two amplified inner cell masses and two trophectoderms from blastocysts.
Project description:We report a filter aided, single tip (FAST) method for ultrasensitive proteomic sample preparation. To minimize sample loss and contamination, the method reduces the surface area of the filter to ~0.1 mm2, the total volume of reagents to < 10 μL, and the number of sample transfer steps to two. 25,883 unique peptides and 3,069 protein groups were identified from 1,000 MCF-7 cells (~100 ng protein content). Single blastomeres from Xenopus laevis embryos at the 50-cell stage (~200 ng yolk free protein/blastomere) generated 20,943 unique peptides and 2,597 protein groups; the proteomic profile clearly differentiated left and right blastomeres, and provides strong support for models in which this asymmetry is established early in the embryo.
Project description:Chromosomal instability (CIN) occurs at high frequency during early in vitro embryogenesis and is known to be associated with early embryonic loss in humans. The chromosomal stability of in vivo-conceived cleavage stage embryos largely remains unknown. Here, we applied haplotyping and copy number profiling to investigate genomic architecture of 171 single bovine blastomeres and to compare the nature and frequency of CIN between in vivo embryos, in vitro embryos produced from ovum pick up with ovarian stimulation (OPU-IVF), and in vitro produced embryos from in vitro matured oocytes without ovarian stimulation (IVM-IVF). Our data shows that CIN is significantly lower in in vivo conceived cleavage stage embryos when compared to in vitro cultured embryos, as genomic stability of single blastomeres in both IVF embryos was severely compromised (P<0.0001)
Project description:In multicellular organisms, heterogametes of oocytes and sperms are fertilized and resulting zygotes give rise to new individuals. The ability of zygotes that produce a fully formed individual from single cell when placed in a supportive environment is defined as totipotency. Given that totipotent cells are the source of all multicellular organisms, better understanding of totipotency has a profound effect on not only biology but also our society. However, the exact distribution of totipotent cells in mammals remains elusive, although zygotes and single blastomeres at 2-cell stage embryos has been thought to be only mouse cells to be totipotent. We now show that a single blastomere isolated from 2- and 4-cell stage embryos gives rise to a fertile adult individual when it placed in a uterus, although isolation of blastomeres at these stage results in the disturbance of transcriptome in single blastomere derived embryos. Single blastomeres from 8-cell and morula stage embryos that were separately cultured in vitro exhibited severe defects in the formation of epiblast and primitive endoderm in the inner cell mass and the development to blastocysts, respectively. Our results thus indicate that totipotency of mouse zygotes extends to single blastomeres of 4-cell stage embryos.
Project description:To determine blastomere fate and embryonic genome activation (EGA) at 5- to 8-cell stage human embryos by global gene expression profile of amplified cDNA from blastomeres at the single cell level
Project description:Whole genome expression analysis reveals that single blastomeres from day-3 human embryos and blastocysts show unique gene signature, thus hESC derived from these two developmental sources might show differential gene expression profile. Comparative gene expression analysis has revealed that no significant differences exist between hESCs derived from blastomeres versus those obtained from ICMs, suggesting that pluripotent hESCs involve a new developmental progression. hESC derived from single blastomeres (VAL-10B and VAL-11B) and from ICM (VAL-5, -7, and -8) were compared by global gene expression
Project description:This study describes the combined sequencing of the genomes and transcriptomes of single blastomeres from mouse 8-cell stage embryos.
Project description:We reported the allele-specific single cell RNA sequencing in highly morphogenic (C57Bl6J-maternal x CASTEiJ-paternal) male F1 mouse embryonic stem cells (mESC) and in vitro mESC derived six cardiac lineage cell types. We demonstrated distinctive gene regulation based on the parental origin of the alleles. We showed that deterministic and stochastic monoalleleic gene classes are distinctive in regulation and are involved in unique processes. In this study we highlighted the importance of parental origin-specific gene expression in development, homeostasis and disease.
Project description:We reported the allele-specific single cell RNA sequencing in highly morphogenic (C57Bl6J-maternal x CASTEiJ-paternal) male F1 mouse embryonic stem cells (mESC) and in vitro mESC derived six cardiac lineage cell types. We demonstrated distinctive gene regulation based on the parental origin of the alleles. We showed that deterministic and stochastic monoalleleic gene classes are distinctive in regulation and are involved in unique processes. In this study we highlighted the importance of parental origin-specific gene expression in development, homeostasis and disease.
