Abstract
Human orphan genes are poorly annotated and lowly expressed, which makes their behaviour in disease difficult to establish. We analysed paired normal and disease RNA-seq libraries from four cohorts, quantified against a combined reference in which 2,190 of 198,507 transcripts are orphan. Cohorts were first screened for library-level differences between the two conditions, and one (GSE40419, lung adenocarcinoma) was excluded because sequencing depth, pseudoalignment rate and submission provenance all separated by condition; it is retained throughout as a calibration. In the remaining three cohorts, comprising 151 matched pairs from psoriasis, laryngeal squamous cell carcinoma and hepatocellular carcinoma, we measured the change in the fraction of transcripts with zero abundance, a quantity that depends on no transformation of expression. Orphan transcripts lose detection relative to non-orphan transcripts matched on abundance and on cross-patient detection frequency, by \( +0.0250 \pm 0.0178 \), \( +0.0553 \pm 0.0073 \) and\( +0.0659 \pm 0.0122 \) once the global shift in detection is held fixed. The three estimates are homogeneous (P = 0.163) and pool to \( +0.0544 \pm 0.0059 \); whereas the same analysis applied to the excluded cohort returns thirteen times that value. We report the result at the level at which it is measured, and withdraw the patient-level and cohort-specific claims made in the first preprint version of this work.