English: Promoter Fragmentation Entropy (PFE)

Epic-seq reasons that cfDNA fragments from active promoters (which are less protected by nucleosomes from endonuclease cleavage) will exhibit more random cleavage patterns than fragments from inactive promoters (which are more protected by nucleosomes from endonuclease cleavage). PFE is a variation of a Shannon Index (a quantitative measure of estimating diversity in a species) used to calculate the diversity of cfDNA fragments’ length where both ends of the fragment fell within the 2 kb flanking each gene’s transcription start site (TSS). The higher the PFE of a gene TSS site, the more likely the gene is highly expressed.

Nucleosome Depleted Region (NDR)

Actively expressed genes have open chromatin at their TSS region, which are less protected by nucleosomes from endonuclease cleavage, thus the depth of cfDNA from active genes’ TSS would be lower than inactive genes’ TSS site. NDR measures the normalized depth within each 2-kilobase (kb) window flanking each TSS (−1,000 to +1,000 bp) in counts per million space. The higher the PFE of a cfDNA from a promotor site, the more likely the gene associated with that cfDNA is highly expressed. The lower the NDR of a gene TSS site, the more likely the gene is highly expressed.