Experiment / E80G0JHGYEpisomal Plasmid MPRA

MPRA stage 1: AD and PSP GWAS variant allelic reporter assay

Functional regulatory variants implicate distinct transcriptional networks in dementia

Both alleles of 5,223 noncoding variants selected from 14 Alzheimer's disease and five progressive supranuclear palsy GWAS loci were tested as 162-bp native-context oligos in an episomal MPRA in HEK293T cells. Six RNA libraries and five plasmid-DNA libraries are represented in the public mapped-count matrix.

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Organism: Human
Taxonomy ID: NCBITaxon:9606
Biosample: CVCL:0063
Reference genome: hg19
Design focus: Variant-focused
Region of interest: Not reported / not applicable

Perturbation & assay details

Basal / Untreated

The library used 162-bp oligos centered on GWAS and high-LD SNVs, with 20-bp random DNA barcodes in the 3' UTR of an eGFP reporter. The oligo was placed upstream of a minimal promoter in a custom plasmid reporter; approximately 8 million HEK293T cells were transfected per replicate with 10 micrograms of library plasmid, harvested 24 hours later, and barcode counts were obtained from plasmid DNA and eGFP mRNA/cDNA. The released matrix contains six HEK293T RNA replicates (HEK1-HEK6) and five plasmid-DNA replicates (Plasmid1-Plasmid5).

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 · 38 definitions

element_id: Stable package identifier for the MPRA1 variant pair.
variant_id: Authors' internal numeric MPRA variant identifier.
rsid: dbSNP rs identifier reported by the authors.
chromosome: Chromosome reported in the author metadata.
position_hg19: Variant coordinate on hg19/GRCh37.
reference_allele: Reference allele (A0) represented by the reference oligo.
alternate_allele: Alternate allele (A1) represented by the alternate oligo.
regulomedb: RegulomeDB annotation supplied by the authors.
construct_type: MPRA1 oligo name orientation: standard, flipped, or mixed when the paired allele names differ.
construct_description: Description of the MPRA1 oligo construction/orientation represented in the library.
ref_oligo_name: Raw reference-allele element name(s) in the GEO count matrix.
alt_oligo_name: Raw alternate-allele element name(s) in the GEO count matrix.
ref_oligo_sequence: Actual 162-bp reference insert recovered from the GEO 210-mer library sequence list.
alt_oligo_sequence: Actual 162-bp alternate insert recovered from the GEO 210-mer library sequence list.
ref_qc_barcode_count: Number of reference barcodes passing the per-barcode count filter.
alt_qc_barcode_count: Number of alternate barcodes passing the per-barcode count filter.
ref_dna_total_raw_count: Sum of raw reference barcode counts across the five plasmid-DNA replicates after barcode QC.
alt_dna_total_raw_count: Sum of raw alternate barcode counts across the five plasmid-DNA replicates after barcode QC.
ref_rna_total_raw_count: Sum of raw reference barcode counts across the six HEK293T RNA replicates after barcode QC.
alt_rna_total_raw_count: Sum of raw alternate barcode counts across the six HEK293T RNA replicates after barcode QC.
ref_dna_mean_raw_count: Mean aggregate reference barcode count per plasmid-DNA replicate.
alt_dna_mean_raw_count: Mean aggregate alternate barcode count per plasmid-DNA replicate.
ref_rna_mean_raw_count: Mean aggregate reference barcode count per RNA replicate.
alt_rna_mean_raw_count: Mean aggregate alternate barcode count per RNA replicate.
ref_dna_mean_cpm: Mean reference plasmid-DNA CPM across DNA replicates after pseudocount normalization.
alt_dna_mean_cpm: Mean alternate plasmid-DNA CPM across DNA replicates after pseudocount normalization.
ref_rna_mean_cpm: Mean reference RNA CPM across HEK293T RNA replicates after pseudocount normalization.
alt_rna_mean_cpm: Mean alternate RNA CPM across HEK293T RNA replicates after pseudocount normalization.
ref_activity_median_log2_rna_dna: Median reference barcode log2 RNA/DNA activity after quantile normalization.
alt_activity_median_log2_rna_dna: Median alternate barcode log2 RNA/DNA activity after quantile normalization.
mpra_log2_effect: Alternate activity minus reference activity in log2 RNA/DNA units.
mpra_p_value: Two-sided tie-corrected normal-approximation Mann-Whitney U p-value for alternate versus reference barcode activity.
mpra_fdr: Benjamini-Hochberg FDR recalculated across QC-passing MPRA1 rows in this package.
mpra_significant_fdr_0_01: Boolean indicating whether mpra_fdr is below 0.01.
mpra_effect_direction: Direction of the alternate-minus-reference activity effect.
rna_replicate_count: Number of HEK293T RNA replicates used (six).
dna_replicate_count: Number of plasmid-DNA replicates used (five).
source_table: Public source count matrix and author variant metadata used for the row.

Quality control

The authors' barcode-level QC was followed: retain a barcode only if it has at least 1 count in every RNA replicate and at least 5 counts in every plasmid-DNA replicate; add a pseudocount of 1, normalize each library to CPM, calculate log2 RNA/DNA ratios using the median plasmid-DNA CPM, and quantile-normalize the six RNA-ratio replicates. Variant pairs with fewer than 5 retained barcodes for either allele were removed. The package retained 4,805 of 5,223 MPRA1 variant pairs from 501,702 eligible barcode rows. Allele activity is the median retained barcode ratio; the p-value is a two-sided Mann-Whitney U test using a tie-corrected normal approximation, and the reported FDR is a Benjamini-Hochberg recalculation across the packaged MPRA1 rows with significance marked at FDR < 0.01.

Curation notes

This is an episomal plasmid MPRA, not a lentiMPRA or STARR-seq experiment. The paper's preprint reports 4,732 MPRA1 variant pairs passing its QC summary, whereas the released GSE163855/GitHub mapped-count files yield 4,805 pairs under the written barcode thresholds; this version/implementation discrepancy is preserved rather than silently forcing the published count. The GEO matrix has five plasmid-DNA columns although the assay used six RNA/biological replicate preparations. MPRA1 raw element names with the _flipped suffix are paired by internal variant ID; actual library insert sequences, including reverse-complement inserts, are retained in the table.

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