TREND enhancer screen for primary human T-cell activation

A pooled synthetic TF-binding-site enhancer library was screened in primary human CD3-positive T cells from two independent donors. Barcode-linked reporter activity was compared between resting cells and cells restimulated through CD3/CD28, yielding donor-resolved activation ratios and a cross-donor summary.

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.

Organism: Human
Taxonomy ID: NCBITaxon:9606
Biosample: CL:0000084
Reference genome: Not reported / not applicable
Design focus: Synthetic / Motif-focused
Region of interest: Not reported / not applicable

Perturbation & assay details

CD3/CD28 activation (24 h restimulation) versus resting state; two independent donors

The same TREND synthetic tandem TF-binding-site library was delivered by lentivirus to primary human CD3-positive T cells. Cells were activated for transduction, cultured, restimulated on day 10 with a CD3/CD28 activator, and collected 24 hours later; resting and stimulated states were assayed by barcode-linked DNA/RNA sequencing. The reporter contains an adenoviral minimal promoter, EGFP, and a 20-bp barcode at the 3-prime end of the EGFP transcript. The table retains the authors' donor-level median RNA/DNA activity scores and stimulation-to-rest ratios, plus arithmetic cross-donor summaries.

Quality control

The authors' pipeline demultiplexed reads, collapsed UMIs, aligned reads to the reference library, removed barcodes with zero DNA or RNA counts, and chose DNA-abundance thresholds intended to retain approximately 50% of constructs in the activation project. The published T-cell analysis used DNA thresholds of 2, 2, 3, and 2 for donor 1 rest, donor 1 stimulated, donor 2 rest, and donor 2 stimulated, respectively, and required at least 8 independent barcodes per enhancer. For this package, rows were retained only when both donors had finite resting activity, stimulated activity, stimulation-to-rest ratio, and barcode counts of at least 8 in both states (8,197 of 57,715 designs). The cross-donor mean columns and direction-consistency flag are derived from the two retained donor records.

Curation notes

The donor identities are intentionally anonymized in the source data. CL:0000084 denotes a T cell, the terminal cell type represented by the primary CD3-positive input population. The source repository also provides a small selected-enhancer XLSX for downstream synthesis; the processed table intentionally represents the full two-donor screen after QC rather than only that selected subset. No allele-contrast variants are present: these are de novo synthetic motif designs.

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TREND enhancer screen for primary human T-cell activation

Processed data

Quality control

Curation notes

Cite OpenMPRA

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