Transcriptome analysis of mouse preimplantation embryos when DNA damage checkpoint is perturbed
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE163765 | GEO | 2021/05/04
REPOSITORIES: GEO
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Project description:Transcriptome analysis of mouse preimplantation embryos when DNA damage checkpoint is perturbed
| PRJNA687487 | ENA
Project description:We systematically analyzed transcriptome in individual human oocytes and preimplantation embryos with a new single-cell RNA seq method we recently developed. We sequenced individual single cells or embryos at seven consecutive stages (mature oocytes, zygotes, 2-cell, 4-cell, 8-cell embryos, morulae, and blastocysts) and single hESC using single-cell RNA seq.
2016-05-13 | E-GEOD-71318 | biostudies-arrayexpress
Project description:The culture conditions of the early preimplantation embryos affects the DNA methylation landscape of the resulting blastocysts.
2015-10-07 | GSE69173 | GEO
Project description:In mammals, chromatin marks at imprinted genes are asymmetrically inherited from each parent to control gene expression. Many genomic imprints are determined by differentially methylated regions (DMRs), but these have not been comprehensively mapped physically or functionally in mouse preimplantation embryos. To address this, we measured genome-wide DNA methylation in individual haploid uniparental parthenogenetic haploid (ph) and androgenetic haploid (ah) E3.5 blastocysts by micro-whole-genome bisulfite sequencing (µWGBS). For comparison, we also included ahESC and phESC lines in the analysis.Comparison of DNA methylomes from uniparental blastocysts with those of control blastocysts (produced by intracytoplasmic sperm injection) identified 859 DMRs. Haploid ES cells showed overall erosion of those DMRs, in contrast to diploid ES cells that relatively maintained differential methylation.
2021-03-02 | GSE152105 | GEO
Project description:40-PE Smart-Seq of RNA isolated from single 2-cell embryos, single morulae, single blastocysts, single E7 embryos epiblast halves and single E7 trophectoderm halves. For all preimplantation stages, embryos were grown ex-vivo from three experimental groups of zygotes: zygotes injected with the mRNA of a recombinant OGA (named BtGH84) (Btgh_injected), zygotes injected with a catalytically dead version of BtGH84 (dBtgh_injected) (control nr.1) and non-injected zygotes (non_injected) (control nr.2). Postimplantation E7 embryos were grown from two experimental group of zygotes: Btgh-injected and dBtgh-injected (control) and transferred at the 2-cell stage to foster mothers. All samples from the same embryonic stage were processed for single embryo (or single-embryo single-tissue) Smart-Seq library preparation at the same time and then sequenced in the same NextSeq500 run. The different embryonic stages were processed on different days and sequenced in different NextSeq500 runs.
2024-03-30 | E-MTAB-12981 | biostudies-arrayexpress
Project description:Study question: Is gene expression in human preimplantation embryos affected by medium used for culturing embryos during an IVF treatment? Summary answer: Six days of in vitro culture of human preimplantation embryos resulted in a medium-dependent expression level of genes involved in apoptosis, protein degradation, metabolism and cell cycle regulation. What is known already: Several human studies have shown an effect of culture medium on embryo development, pregnancy outcome and birthweight. However, the underlying mechanisms in human embryos are still unknown. In animal blastocysts, it has been demonstrated that culture of preimplantation embryos affects gene expression. In humans, it has been found that culture medium affects gene expression of cryopreserved embryos that, after thawing, were cultured in two different media for 2 more days. Study design, size, duration: In a multicenter trial, women were randomly assigned to two culture media groups (G5 and HTF). Data on embryonic development were collected for all embryos. In one center, embryos originating from 2PN zygotes that were not selected for transfer or cryopreservation on day 2 or 3, were further cultured until day 6 and collected for this study, if couples consented. Participants/materials, setting, methods: Ten blastocysts from each study group, matched for fertilization method, maternal age and blastocyst quality, were selected and their mRNA was isolated and amplified. Embryos were individually examined for genome-wide gene expression using Agilent microarrays and PathVisio was used to identify the pathways that showed a culture medium-dependent activity. Main results and the role of chance: Expression of 951 genes differed significantly (P < 0.01) between the two study groups. Eighteen pathways, involved in apoptosis, metabolism, protein processing and cell cycle regulation, showed a significant overrepresentation of differentially expressed genes (DEGs). The DNA replication, G1 to S cell cycle control and oxidative phosphorylation pathways were upregulated in the G5 group compared with the HTF group. This is in agreement with the higher number of cells seen in G5 embryos on day 2 and 3. Limitations, reasons for caution: Despite careful matching of the embryos, it cannot be ruled out that differences observed between the study groups are affected by factors not investigated. Extrapolation of these results to embryos used for transfer demands caution as in the present study embryos that were not selected for either embryo transfer or cryopreservation have been used for the culture experiment until day 6. Wider implications of the findings: This study shows that gene expression in human preimplantation embryos is affected by the culture medium used during an IVF treatment. Furthermore, it provides insight into the biological mechanisms that are affected. The increased cell cycle regulation is reflected by a higher number of blastomeres. Whether these results are also connected to long-term effects remains unknown. However, it is not unlikely that early adaptations of the preimplantation embryo to its environment persist during fetal and post-natal development. Note: this study has been conducted using the Agilent-028004 SurePrint G3 Human GE 4x180K Microarray. This array contains the same reporters as the Agilent-028004 SurePrint G3 Human GE 8x60K Microarray. As such we have linked this entry to the A-GEOD-14550 array design, belonging to the latter.
2015-07-08 | E-MTAB-3405 | biostudies-arrayexpress
Project description:Age of the donor animal has a significant influence on the oocyteM-bM-^@M-^Ys ability to complete maturation and acquire the mRNA and proteins required for normal embryonic development. It is well known that developmental competence is reduced in oocytes derived from juvenile animals when compared to that derived from adults. However, the molecular mechanisms associated with these differences are not well elucidated. The aim of this study was to analyze the transcriptome differences between adult versus prepubertal derived preimplantation embryos and to identify genes associated with oocyte developmental potential. We performed cDNA microarray analysis on populations of preimplantation embryos (8- to 16-cell and Day 7 blastocysts) derived from adult versus prepubertal Japanese Black cattle. Total RNA was extracted and amplified in a linear fashion and then subjected to the microarray hybridization using the Affymetrix GeneChip Bovine Genome Array. The Bovine Array contains 24 072 probe sets representing over 23000 transcripts and 19000 UniGene clusters. Following normalization of the microarray data, analysis revealed differences in gene expression abundance in 8- to 16-cell and blastocysts harvested from adult versus prepubertal heifers. Results indicated that a total of 591 and 490 genes were differentially expressed (M-bM-^IM-%2-fold) in 8- to 16-cell and blastocyst respectively between the two groups (p<0.01). In 8- to 16-cell stage, 373 genes were up-regulated and 218 gens down-regulated in adult when compared to the prepubertal heifer group. In case of blastocyst stage embryos, 242 genes were up-regulated and 248 genes were down-regulated in adult when compared to prepubertal group. The ovaries of adult cows (Japanese black cattle) were collected from local abattoir while ovaries of prepubertal Japanese black heifers (9 to 12 months old) were collected by spay device at several commercial farms. The ovaries were transported to the research laboratory in 0.67% (w/v) NaCl solution containing 100 mg/L kanamycin sulfate. COC were placed in 200 M-BM-5L drop of the maturation medium (TCM 199 + 10% FBS) and cultured at 38.5M-BM-0C in a humidified atmosphere of 5% CO2 in air for 20 to 22 h for in vitro maturation (IVM). After 20 to 22 h IVM, the COC were fertilized in vitro with frozenM-bM-^@M-^Sthawed semen of Japanese Black bull in BO medium. Six hours after IVF, oocytes were denuded of surrounding cumulus cells by gentle pipetting in a solution of 0.1% (w/v) hyaluronidase. The presumptive zygotes were cultured in CR1aa medium supplemented with fatty acid-free BSA (Sigma) for 2 days and cultured in CR1aa medium supplemented with 10% (v/v) fetal bovine serum (FBS) for next 5 days at 38.5 M-BM-0C in 5% CO2, 5% O2 and 90% N2 with high humidity (15 to 20 zygotes per 50 M-BM-5l microdrop). 8- to 16-cell refers to day 3, 70 to 72 hours post insemination( hpi) preimplantation embryo stage containing 8 to 16 cells, while blastocyst refer to pre-hatching day 7 expanded blastocysts. Pools of 8- to 16- cell embryos (50 in each sample) and blastocysts (10 in each sample) from prepubertal and adult groups were used for cDNA microarray.
2012-05-31 | E-GEOD-24596 | biostudies-arrayexpress
Project description:DNA demethylation of paternal genome in zygotes takes place in various mammals including mice and human. Recent studies have revealed that this is achieved through Tet3-mediated iterative oxidation of 5-methylcytosine (5mC) coupled with replication-dependent dilution. Tet3-mediated paternal DNA demethylation is believed to be required for mouse development given that Tet3 heterozygous embryos, derived by fertilizing Tet3 knockout (KO) oocytes with wild-type (WT) sperms, exhibit 5mC oxidation defects and embryonic sublethality, Here we demonstrate that the sublethality phenotype of the maternal KO mice is caused by haploinsufficiency of Tet3, but not by defective paternal 5mC oxidation. We found that Tet3 heterozygous mice derived from crosses of heterozygous father or mother with WT mice also exhibit sublethality phenotype similarly to Tet3 maternal KO mice. Importantly, embryos reconstituted with WT paternal nuclei that bypassed 5mC oxidation develop to term and grow to adulthood normally. Genome-scale DNA methylation analysis of the maternal KO zygotes and blastocysts demonstrated that hypermethylation caused by the depletion of maternal Tet3 is largely diminished by the blastocyst stage. Our study thus not only demonstrates that Tet3-mediated paternal 5mC oxidation is dispensable for mouse development but also suggests the existence of a compensation mechanism in preimplantation embryos that can compensate for the defective 5mC oxidation in zygotes. This data set includes RRBS data of wild-type and maternal Tet3 KO zygotes and blastocysts (C57BL/6J x CAST/EiJ)
2015-02-28 | E-GEOD-62719 | biostudies-arrayexpress
Project description:Germline genome editing by the CRISPR/Cas9 system has potential for generation of genetically modified animals/the targeted correction of pathological mutations, understanding the mechanism of early embryonic development. Previous reports showed that Cas9 protein without sgRNAs could induce genomic damage in cell levels in vitro. However, whether the detrimental effects occurred in embryos after Cas9-treatment remains unknown. Here, by using pig embryos as the subjective, we found that Cas9 protein transcribed from injected Cas9 mRNA could persist to at least blastocyst stage. Cas9 protein alone could induce the genome damage of preimplantation embryos, represented by increased phosphorylated histone H2AX (pH2AX) foci on the chromatin fiber, leading to apoptosis and decreased cell number of blastocysts. In addition, single-blastocyst RNA sequencing confirmed that Cas9 protein without sgRNAs could also cause the blastocyst transcriptome changes, depressing the embryo development signal pathway, such as cell cycle, metabolism, and cellular communication related signal pathways, while activating apoptosis and necroptosis signal pathway, both of which together resulted in impaired the preimplantation embryonic development.
2023-02-05 | GSE224238 | GEO
Project description:The G2 DNA damage checkpoint inhibits Cdc2 and mitotic entry through the dual regulation of Wee1 and Cdc25 by the Chk1 effector kinase. Up-regulation of Chk1 by mutation or overexpression bypasses the requirement for up-stream regulators or DNA damage to promote a G2 cell cycle arrest. We screened in fission yeast for mutations that rendered cells resistant to overexpressed Chk1. We identified a mutation in tra1, which encodes one of two homologs of TRRAP, an ATM/R-related pseudokinase that scaffolds several histone acetyltransferase (HAT) complexes. Inhibition of histone deacetylases reverts the resistance to overexpressed Chk1, suggesting this phenotype is due to a HAT activity, though expression of checkpoint and cell cycle genes is not greatly affected. Cells with mutant or deleted tra1 activate Chk1 normally and are checkpoint proficient. However, these cells are semi-wee even when overexpressing Chk1, and accumulate inactive Wee1 protein. The Cdr (changed division response) kinases Cdr1 and Cdr2 are negative regulators of Wee1, and while best characterized in the cellular response to limited nutrition, we show that they are required for the Tra1-dependent alterations to Wee1 function. This identifies Tra1 as another component controlling the timing of entry into mitosis via Cdc2 activation. Two and three independent microaaray experiments were done for wild type and the mutant (tra1-1) respectively.
2010-04-11 | E-GEOD-12674 | biostudies-arrayexpress