Removal of unwanted variation in pseudobulk analysis of single-cell RNA sequencing data and the leveraging of pseudoreplicates.

ABSTRACT: Removing unwanted variation (RUV) is key for accurate biological interpretation in high-throughput sequencing studies. However, no standardized approach exists for pseudobulked single-cell RNA-sequencing (scRNA-seq) data. Improper implementation of RUV methods may remove biological information, jeopardizing power and false positive control in differential expression analysis. We evaluate the impact of three implementation strategies ('trails') in three RUV methods (RUV2, RUVIII, RUV4) using simulated and real biological signals in pseudobulked scRNA-seq data. Effects of technical noise under confounding and model misspecification conditions are also considered. Additionally, we introduce a novel strategy, RUVIII PBPS, to remove unwanted variation in pseudobulk differential expression analy

SUBMITTER: Prieto Leon S 

PROVIDER: S-EPMC12684399 | biostudies-literature | 2025 Dec

REPOSITORIES: biostudies-literature