Allele-specific STARR-seq of AMD-associated SNPs in ARPE-19
Single-cell multiome and enhancer connectome of human retinal pigment epithelium and choroid nominate causal variants in macular degenerationA focused allele-specific STARR-seq library tested reference and alternative alleles centered in 152 bp of genomic context for AMD-associated SNPs and MAPT-locus control SNPs in the human RPE cell line ARPE-19. The same input plasmid library was measured after 24 hours in four biological replicates of naïve culture and four biological replicates of complement-activated culture; the table reports the final QC-passing variant-level enhancer and allelic activity results for both conditions.
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
Naïve ARPE-19 culture with 10% FBS versus complement-activated culture with 10% complement-competent human serum for 18 h; shared input plasmid library
Reference and alternative alleles for each SNP were synthesized in 152 bp of genomic context, each with a 10-nucleotide degenerate barcode, and cloned into the hSTARR-seq_ORI vector (Addgene #99296) downstream of a minimal promoter. Ten micrograms of plasmid library per 10 million ARPE-19 cells were delivered by Lonza Nucleofector. Reporter RNA was poly(A)-selected, DNase-treated, reverse-transcribed with a target-specific primer, PCR-amplified, and sequenced as paired-end 150-bp reads; input plasmid DNA was sequenced in parallel. Four biological replicates were generated for each output condition. Barcode counts were normalized with DESeq2, enhancer activity was called against the input library, and alternative-versus-reference allele effects were estimated with mpralm from the mpra package.
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 · 16 definitions
- variant_set
- Variant class: AMD-risk for disease-associated SNPs or MAPT-control for random control SNPs.
- variant_id
- dbSNP rs identifier for the tested SNP.
- chromosome
- Chromosome reported by the authors in the hg38 coordinate system.
- position_grch38
- SNP position reported in hg38/GRCh38 coordinates.
- reference_allele
- Reference allele represented by the reference oligonucleotide.
- alternate_allele
- Alternative allele represented by the alternative oligonucleotide.
- source_sheet
- Data S11 workbook sheet from which the row was taken: AMD SNPs or Control SNPs.
- naive_in_starr_enhancer
- Boolean indicating whether the SNP oligonucleotide was called a STARR-seq enhancer in naïve ARPE-19 culture.
- naive_log2fc_alt_vs_ref
- mpralm allele-specific log2 fold change of alternative versus reference activity in naïve ARPE-19 culture.
- naive_adjusted_p_value
- Multiple-testing-adjusted p-value for the alternative-versus-reference effect in naïve culture.
- naive_functional_snp
- Author-defined functional-SNP flag in naïve culture; blank for MAPT controls because the control sheet does not report this classification.
- complement_in_starr_enhancer
- Boolean indicating whether the SNP oligonucleotide was called a STARR-seq enhancer in complement-activated ARPE-19 culture.
- complement_log2fc_alt_vs_ref
- mpralm allele-specific log2 fold change of alternative versus reference activity in complement-activated culture.
- complement_adjusted_p_value
- Multiple-testing-adjusted p-value for the alternative-versus-reference effect in complement-activated culture.
- complement_functional_snp
- Author-defined functional-SNP flag in complement-activated culture; blank for MAPT controls because the control sheet does not report this classification.
- complement_minus_naive_log2fc
- Descriptive difference between complement-activated and naïve alternative-versus-reference log2 fold changes; not an independently tested interaction statistic.
Quality control
The authors merged paired-end reads, retained primary alignments with the expected adapter and exactly 10-bp barcodes, and discarded oligonucleotides with fewer than 5 unique barcodes in any replicate. Unique-barcode counts were normalized with DESeq2; enhancer calls required absolute fold change >1.5 and adjusted p <0.05 relative to the input library. Allele-specific effects were calculated with mpralm, and functional SNPs required enhancer membership, adjusted p <0.05, and an absolute allele effect above the 80th percentile of MAPT control SNP effects. The paper reports same-condition replicate correlations of 0.972–0.994 and final analysis of 1,450 AMD SNPs after QC. Package QC removed blank trailing rows and retained only rows with a variant ID, valid hg38 coordinate and nucleotide alleles, and finite adjusted p-values and effect estimates, yielding 1,450 AMD-risk and 357 MAPT-control SNP rows; nonsignificant variants were retained.
Curation notes
The raw GEO barcode matrix is retained as deposited and contains 11,912 allele-oligo rows with several namespace prefixes (including AMD, AMDRE, RE, PACG, and HM), some of which are not represented in the final Data S11 tables. To avoid misassigning counts across those namespaces, the processed table uses the publication's final Data S11 mpralm results and retains the barcode matrix separately for provenance. Raw SRA/FASTQ reads were not downloaded because the study's sequencing data are large and the user requested MPRA-relevant data rather than raw sequence reads. ARPE-19 resolves to Cellosaurus CVCL:0145.