Experiment / E3CDDUF4GEpisomal Plasmid MPRA

Nanog locus Upstream combinatorial MPRA

Systematic analysis of intrinsic enhancer-promoter compatibility in the mouse genome

An episomal Upstream combinatorial MPRA library tested pairwise combinations of approximately 450-bp DNase-hypersensitive candidate CREs and GENCODE-annotated promoter fragments from the Nanog genomic locus. The clean table preserves both cloned orientations and reports normalized reporter activity and corrected cCRE-promoter boost indices across three biological replicates in E14TG2a mouse embryonic stem cells.

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Organism: Mouse
Taxonomy ID: NCBITaxon:10090
Biosample: CVCL:9108
Reference genome: mm10
Design focus: Region-focused
Region of interest: chr6:122308357-123303928

Perturbation & assay details

Basal / Untreated (E14TG2a mESCs cultured in 2i+LIF)

cCREs and promoters were PCR-amplified from E14TG2a genomic DNA, pooled, randomly ligated into duplets, and cloned upstream of a randomly barcoded promoterless transcription unit. The library therefore contains cCRE-promoter, cCRE-cCRE, promoter-promoter, and promoter-cCRE pairs; this child experiment contains the cCRE-promoter subset, retaining both orientations. Three biological replicates were transiently transfected, barcodes were counted in cDNA and plasmid DNA, and activity was summarized as the geometric mean of normalized cDNA:pDNA ratios. Klf2 and Nanog Upstream libraries were mixed for transfection.

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 · 32 definitions

combination_id: Published cCRE-promoter pair identifier including cloned orientations.
combination_class: Source combination class; every retained row is EP, a cCRE-promoter pair.
enhancer_id: Study identifier for the cCRE in the first reporter position.
enhancer_orientation: Orientation of the cCRE in the reporter construct (+ or -).
enhancer_chrom: mm10 chromosome for the cCRE fragment.
enhancer_start_mm10: 0-based half-open mm10 start coordinate of the cCRE fragment.
enhancer_end_mm10: 0-based half-open mm10 end coordinate of the cCRE fragment; end-start is 450 bp.
enhancer_sequence_mm10_forward: Reference forward-strand sequence of the cCRE insert from Data S1; it is not reverse-complemented when the cloned orientation is -.
promoter_id: Study identifier for the promoter in the second reporter position.
promoter_orientation: Orientation of the promoter in the reporter construct (+ or -).
promoter_chrom: mm10 chromosome for the promoter fragment.
promoter_start_mm10: 0-based half-open mm10 start coordinate of the promoter fragment.
promoter_end_mm10: 0-based half-open mm10 end coordinate of the promoter fragment; end-start is 450 bp.
promoter_sequence_mm10_forward: Reference forward-strand sequence of the promoter insert from Data S1; it is not reverse-complemented when the cloned orientation is -.
fragment_distance_bp: Distance in base pairs as deposited in the GEO source field dist.
pair_activity_rep1: Normalized cDNA:pDNA reporter activity for biological replicate 1.
pair_activity_rep2: Normalized cDNA:pDNA reporter activity for biological replicate 2.
pair_activity_rep3: Normalized cDNA:pDNA reporter activity for biological replicate 3.
pair_activity_geometric_mean: Geometric mean pair activity across the three biological replicates, as deposited in activity_all.
enhancer_single_activity_rep1: Normalized activity for the cCRE fragment alone in the first position, from activity_br1_frag1.
enhancer_single_activity_rep2: Normalized activity for the cCRE fragment alone in the first position, from activity_br2_frag1.
enhancer_single_activity_rep3: Normalized activity for the cCRE fragment alone in the first position, from activity_br3_frag1.
enhancer_single_activity_geometric_mean: Geometric mean cCRE singleton activity across three replicates, from activity_all_frag1.
promoter_single_activity_rep1: Normalized activity for the promoter fragment alone in the second position, from activity_br1_frag2.
promoter_single_activity_rep2: Normalized activity for the promoter fragment alone in the second position, from activity_br2_frag2.
promoter_single_activity_rep3: Normalized activity for the promoter fragment alone in the second position, from activity_br3_frag2.
promoter_single_activity_geometric_mean: Geometric mean promoter singleton activity across three replicates, from activity_all_frag2.
promoter_baseline_median_activity: Deposited promoter-specific baseline median used in the source boost-index processing; blank where Data S3 reports NA.
promoter_baseline_n: Deposited number of baseline observations used for the source baseline; blank where Data S3 reports NA.
boost_index_log2: Corrected log2 cCRE-promoter boost index from Data S3/GEO, measuring pair activity relative to the promoter-specific baseline.
locus: Study locus label; Nanog for every row in this child experiment.
source_locus_pair_id: Deposited idlocus identifier combining the pair and locus.

Quality control

The authors aligned inverse-PCR reads to a custom fragment genome, clustered barcode sequencing errors with Starcode, removed barcodes present in multiple libraries or assigned to multiple element combinations, normalized cDNA and pDNA counts per sample, required at least 5 barcodes per singlet or combination and at least 8 pDNA counts per barcode, and calculated geometric-mean activities across three biological replicates. Upstream replicate concordance was reported as mean Pearson r=0.87 (range 0.83-0.90). Package QC restricted the deposited Data S3/GEO table to class=EP, required nonempty element identifiers and valid mm10 coordinates, and retained only finite three-replicate/aggregate pair activities and finite BoostIndex; 1,256 of 1,321 Nanog EP rows passed, with 65 -Inf BoostIndex rows excluded. Auxiliary fields remain blank where the source reports NA.

Curation notes

The source Data S3/GEO table contained 1,321 Nanog EP records; 1,256 finite-score records are packaged. The source reports NA for Basal_median and Nbasal for all Nanog rows, so those columns are intentionally blank even when BoostIndex is finite. The table keeps orientation-specific pairs; the article averages cCRE orientations for several downstream analyses. Data S1 sequences were joined by fragment identifier and locus. This is a published processed-results table, not a raw barcode-count matrix.

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