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. 2020 Apr 1;12(5):e12112. doi: 10.15252/emmm.202012112

Figure 1. Development of MPRA for use in primary human CD4 T cells.

Figure 1

  • A
    Experimental workflow for MPRA: oligonucleotide library is cloned into an empty vector, and a reporter gene and promoter are inserted using restriction sites within the oligonucleotide. The assembled plasmid is transfected into primary CD4 T cells, and RNA is extracted after 24 h. RNA barcode counts are normalised to their respective counts in the input plasmid library (DNA), which is sequenced separately.
  • B
    Adapted MPRA plasmid incorporating RSV promoter.
  • C
    Principal component analysis of scaled element counts (sum of barcodes tagging same genomic construct in mRNA) in resting and stimulated CD4 T cells from 12 donors. Dotted lines indicate samples from the same donor.
  • D
    Heat maps showing pairwise comparison of MPRA activity for all constructs (mRNA/DNA) between donors—left panel: resting CD4 T cells; right panel: stimulated CD4 T cells.