Fig. 1

Study overview and RNA-seq analysis pipeline. Forty-one participants performed twelve weeks of resistance training with low- (one set per exercise, LOW) and moderate-volume (three sets per exercise, MOD) in a contralateral manner (2–3 sessions week-1) (a). Pre- and post-training testing included strength and muscle lean-mass assessments. Muscle biopsies were collected from m. vastus lateralis at four time-points, prior to and after the intervention (Week 0 and 12) and before and after the fifth training session (Week 2). Biopsies from participants who completed > 85% of prescribed sessions were used for RNA extraction (n = 34; A). RNA quality was assessed (b), and participants with RNA quality indicator (RQI) scores > 7 were submitted for RNA-seq (n = 25). RNA quality was not associated with muscle tissue weight (c), and participants included in RNA-seq experiments did not differ from excluded in terms of limb lean-mass gains (d). Higher training volume led to greater gains in limb lean mass (e) and strength (f) in the lower extremities (n = 25). RNA-seq data were quality filtered using trimgalore and trimmomatic and reads were compared to unfiltered reads (g). Read alignment was performed using five tools of which RSEM, kallisto, and Salmon showed greater fractions of genes with robust expression after removing low-abundance genes (expression filtering; H) compared to HISAT2 and STAR. RSEM, kallisto and Salmon also showed less Log2-differences between biological replicates in a subset of genes with known robust expression (see text for details, i)