Project description:In total, 240 single blastomeres from nine top-quality day-4 embryos frozen at day 3 of development and four fresh top-quality day-4 embryos that had one-cell biopsy on day 3 for preimplantation genetic diagnosis (PGD) were collected. Blastomeres' DNA was amplified using SurePlex DNA Amplification System (BlueGnome, Cambridge, UK) . Array-CGH was carried out using 24Sure Cytochip microarrays following the standard protocol (BlueGnome, www.cytochip.com).
Project description:In total, 240 single blastomeres from nine top-quality day-4 embryos frozen at day 3 of development and four fresh top-quality day-4 embryos that had one-cell biopsy on day 3 for preimplantation genetic diagnosis (PGD) were collected. Blastomeres' DNA was amplified using SurePlex DNA Amplification System (BlueGnome, Cambridge, UK) . Array-CGH was carried out using 24Sure Cytochip microarrays following the standard protocol (BlueGnome, www.cytochip.com). BAC array-CGH on single blastomeres amplified by SurePlex amplification Kit.
Project description:Seventy blastomeres from fourteen frozen-thawed supernumerary human preimplantation embryos were disassociated and genomic was amplified using Multiple Displacement Amplification. BAC array-CGH was performed on the amplified products. BAC array-CGH on single blastomeres amplified by Multiple Displacement Amplification.
Project description:Seventy blastomeres from fourteen frozen-thawed supernumerary human preimplantation embryos were disassociated and genomic was amplified using Multiple Displacement Amplification. BAC array-CGH was performed on the amplified products.
Project description:In this study, we extend array CGH technology by making the accurate detection of chromosomal imbalances possible from a single fibroblast and blastomere following Phi29 DNA polymerase amplification. Firstly, array CGH experiments were performed on four different fibroblast cell lines, derived from patients affected by, respectively, trisomy 13, 18, 21, and monosomy X. For each cell line, three single cells were amplified. Following DNA amplification, all cells showed the expected DNA yields (n=12; 1.87 µg plus:minus 0.39). Sex-mismatch array CGH experiments were conducted on amplified DNA samples obtained from each cell. Sex chromosome ploidy levels, as well as all expected autosomal abnormalities were clearly identified. Secondly, we applied single-cell aneuploidy screening for the detection of chromosomal imbalances in preimplantation embryos. DNA from blastomeres from three 7-8 cell-stage embryos was amplified by ï¦29 DNA polymerase. Following DNA amplification, all cells yielded the expected amount of DNA (n=16; 2.45 µg plus:minus 0.41). Chromosomal aneuploidies were accurately detected using a simple and rapid array CGH protocol.
Project description:Detection of genomic rearrangements from a single cell instead of a population of cells is an emerging research technique with important applications in the study of human fertility, constitutional chromosomal disorders, and tumor progression. Here, we develop a method to improve the detection of single-cell genome-wide copy number variation. Additional information about the blastomeres can be found in GSE11663. At this study, 14 amplified single blastomere DNA samples derived from 3-day-old and 4-day-old human embryos were analyzed by Agilent 244K array CGH. For these single cell Agilent 244K array CGH analyses: non-amplified genomic DNA extracted from the blood of a Klinefelter patient (XXY) was used as a reference sample. As a validation, the corresponding non-amplified genomic DNA samples were analyzed by 250K Nsp I SNP arrays (platform GPL3718 and GSE11663).
Project description:Embryonic chromosome aberrations cause birth defects and reduce human fertility. However, neither their nature nor incidence are known. Here, we develop a method to assess genome-wide copy number variation and loss of heterozygosity in single cells and apply it to screen blastomeres from in vitro fertilized preimplantation embryos. Complex patterns of chromosome-arm imbalances or segmental deletions, duplications or amplifications that were reciprocal in sister blastomeres were detected in a large proportion of the embryos. In addition, aneuploidies and uniparental isodisomies were frequently observed. Since these embryos were derived from young fertile couples, the data indicate that chromosomal instability is common to human embryogenesis. Keywords: comparative genomic hybridisation
Project description:Despite intensive efforts, establishing porcine embryonic stem cells have been challenging. We recently derived mouse expanded potential stem cells (EPSCs) from individual blastomeres by inhibiting the activity of critical molecular pathways that predisposes lineage differentiation in the mouse preimplantation embryo. EPSCs had enriched molecular signatures of blastomeres and possessed the developmental potency to all embryonic and extraembryonic cell lineages. In this study, we report the derivation of porcine EPSC (pEPSC) lines either directly from preimplantation embryos or by reprogramming fetal fibroblasts. Under similar culture conditions, human ESCs and iPSCs can be converted, or somatic cells are directly reprogrammed, to EPSCs (hEPSCs) that display the molecular and functional attributes reminiscent of pEPSCs. Here, we performed Single-Cell RNA-seq experiments to characterise the transcriptional heterogeneity of the EPSCs.