Gene Expression
Nebulas
Gene Expression NebulasSummary: The study of early post-implantation human embryo development is limited by the scarcity of embryo samples and ethical restrictions. We developed a human pluripotent stem cell (hPSC)-based microfluidic platform to model several developmental processes during this stage. We applied single-cell mRNA sequencing technique to examine the transcriptomes of posteriorized embryonic-like sacs obtained using the microfluidic platform, as well as of amniotic ectoderm-like cells generated using a Transwell method and human embryonic stem cells (H9 hESCs).
Overall Design: Cells retrieved from posteriorized embryonic-like sacs were pooled from 6 microfluidic devices, after 48 hours of exogenous BMP4 stimulation. Amniotic ectoderm-like cells were also obtained by treating H9 hESCs with BMP4 for 48 hours using a Transwell method. H9 hESCs maintained using standard tissue culture dishes were also sequenced for comparison.
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| Growth Protocol: | - |
| Treatment Protocol: | - |
| Extract Protocol: | Posteriorized embryonic-like sacs (48 hours) within the microfluidic devices, amniotic-like cells derived using transwell and H9 cells were dissociated into single cells by incubating in Accutase for 1 hour. Cells from posteriorized embryonic-like sacs were lysed and barcoded with 10x Chromium Controller Instrument (10x Genomics). Amniotic-like cells derived using transwell and H9 cells were mixed at a 2:1 ratio, before lysed and barcoded with 10x Chromium Controller Instrument. |
| Library Construction Protocol: | cDNA library was constructed according to the standard 10x Genomics scRNA-seq library construction protocol (Single Cell 3' v2). The libraries were sequenced on Illumina HiSeq 4000 with paired-end sequencing (One sample per flow cell). |
| Molecule Type: | poly(A)+ RNA |
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| Library Layout: | PAIRED |
| Library Strand: | Forward |
| Platform: | ILLUMINA |
| Instrument Model: | Illumina HiSeq 4000 |
| Strand-Specific: | Specific |
| Data Resource | GEN Sample ID | GEN Dataset ID | Project ID | BioProject ID | Sample ID | Sample Name | BioSample ID | Sample Accession | Experiment Accession | Release Date | Submission Date | Update Date | Species | Race | Ethnicity | Age | Age Unit | Gender | Source Name | Tissue | Cell Type | Cell Subtype | Cell Line | Disease | Disease State | Development Stage | Mutation | Phenotype | Case Detail | Control Detail | Growth Protocol | Treatment Protocol | Extract Protocol | Library Construction Protocol | Molecule Type | Library Layout | Strand-Specific | Library Strand | Spike-In | Strategy | Platform | Instrument Model | Cell Number | Reads Number | Gbases | AvgSpotLen1 | AvgSpotLen2 | Uniq Mapping Rate | Multiple Mapping Rate | Coverage Rate |
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