Individual Oct4-bound enhancer STARR-reporter assay in E14 mouse ESCs
Pluripotency reprogramming by competent and incompetent POU factors uncovers temporal dependency for Oct4 and Sox2An episomal STARR-reporter/qPCR assay tested two Oct4-bound mouse genomic regions near Sox21 and Spata13, each with a site-directed motif mutant, alongside a CMV positive control and three glucocorticoid-receptor negative controls. E14 mouse embryonic stem cells were transiently transfected with individual constructs, harvested after 24 hours, and GFP reporter expression was measured in four biological replicates.
Processed tables are specific to each experiment. Column names, units, measurements, and table structure are not standardized across the database. Check this experiment’s column definitions and quality-control notes before comparing or combining data.
Perturbation & assay details
Basal / Untreated (ESC medium with 20% FBS and LIF)
Individual low-complexity validation using the episomal pSTARR-seq_human reporter vector (Addgene #71509). The tested inserts were two nested-PCR genomic fragments and their site-directed motif mutants, plus gBlock controls; E14 mESCs were plated at 2.5 × 10^4 cells per well, transfected with 1 µg plasmid using Lipofectamine 2000, and collected 24 h later. Reporter GFP RNA was quantified by RT-qPCR normalized to Rpl19. This arrayed/qPCR STARR validation is distinct from the paper's separate genome-wide FAIRE-STARR-seq library analysis.
Processed data
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Filters apply to this table only. The CSV download contains the complete processed table; filtered rows are available through the API.
Column dictionary · 24 definitions
- construct_id
- Unique source construct name from Supplementary Table 8 and Source Data Figure 2J.
- construct_class
- Functional class assigned from the source labels: positive control, negative control, test enhancer, or motif mutant.
- source_sample_type
- Original sample-type label from Source Data Figure 2J.
- target_gene
- Nearby gene for the genomic enhancer fragment; NA for synthetic controls.
- genomic_location_mm10
- Broad mm10 interval reported for the genomic fragment; NA for synthetic controls.
- sequence_5prime_to_3prime
- 5′ to 3′ DNA sequence of the reporter insert from Supplementary Table 8.
- sequence_length_bp
- Number of bases in the tested reporter insert sequence.
- motif
- Regulatory motif or control motif represented by the construct.
- motif_state
- Reference, mutated, positive-control, scrambled-control, or synthetic-control state.
- generation_method
- How the insert was generated: gBlock, nested PCR, or site-directed mutagenesis.
- expression_exp1
- Source GFP/Rpl19-normalized reporter expression for biological replicate Exp1.
- expression_exp2
- Source GFP/Rpl19-normalized reporter expression for biological replicate Exp2.
- expression_exp3
- Source GFP/Rpl19-normalized reporter expression for biological replicate Exp3.
- expression_exp4
- Source GFP/Rpl19-normalized reporter expression for biological replicate Exp4.
- mean_expression
- Source mean reporter expression across the four biological replicates.
- sd_expression
- Source sample standard deviation of reporter expression across the four biological replicates.
- negative_control_mean_expression
- Derived mean of the three source negative-control construct means, repeated as the assay baseline.
- activity_fold_vs_negative_controls
- Derived mean_expression divided by the three-negative-control baseline.
- log2_activity_vs_negative_controls
- Derived log2 activity fold relative to the three-negative-control baseline.
- parent_construct
- Matched reference construct for a motif mutant; NA for non-mutant rows.
- parent_mean_expression
- Mean expression of the matched parent construct; NA when not applicable.
- mutant_vs_parent_log2
- Derived log2 ratio of mutant mean expression to its matched parent mean; NA for non-mutant rows.
- source_table
- Publisher source locations for sequence and quantitative fields.
- qc_status
- Package-level retention status; PASS indicates the finite/nonnegative replicate and summary checks passed.
Quality control
The authors used three glucocorticoid-receptor enhancer controls as negative controls for basal promoter activity, a CMV enhancer positive control, four biological replicates (Exp1–Exp4), and Student's t-tests for Figure 2J comparisons. Package QC retained all eight constructs because every replicate expression value and source mean/standard deviation was finite and nonnegative; no rows were excluded or imputed. The derived negative-control baseline is the arithmetic mean of the three source negative-control means (1.458333), activity_fold_vs_negative_controls is mean_expression divided by that baseline, and mutant_vs_parent_log2 is log2(mutant mean / matched wild-type mean).
Curation notes
The paper also reports a separate genome-wide FAIRE-STARR-seq assay with three biological replicates and MACS2 enhancer calls, but its underlying STARR input/output count or peak tables are not deposited in the paper's GEO series. GSE103980 contains the paper's ChIP-seq, RNA-seq, and ATAC-seq subseries only, so the child package represents only the directly available Figure 2J STARR-reporter measurements. Coordinates and sequences are transcribed from Supplementary Table 8; quantitative values are from the Figure 2J sheet of the Source Data workbook. The methods specify generic E14 mouse ESCs without a catalog or subclone, so biosample_id uses the generic ES-E14 Cellosaurus record CVCL:C320; the related E14TG2a record CVCL:9108 is not asserted.