The Effect of Human Genome Annotation Complexity on RNA-Seq Gene Expression Quantification

Next-generation sequencing (NGS) has brought human genomic research to an unprecedented era. RNA-Seq is a branch of NGS that can be used to quantify gene expression and depends on accurate annotation of the human genome (i.e., the definition of genes and all of their variants or isoforms). Multiple annotations of the human genome exist with varying complexity. However, it is not clear how the choice of genome annotation influences RNA-Seq gene expression quantification. We assess the effect of different genome annotations in terms of (1) mapping quality, (2) quantification variation, (3) quantification accuracy (i.e., by comparing to qRT-PCR data), and (4) the concordance of detecting differentially expressed genes. External validation with qRT-PCR suggests that more complex genome annotations result in higher quantification variation.