RNA extraction, sequencing, and data processing
DNA and RNA were extracted from chest skin biopsies using TRIzol™
Reagent and the Zymo Quick-DNA/RNA™ Microprep Plus Kit (Zymo Research,
Irvine, CA). We quantified RNA integrity (RQN) using a Fragment Analyzer
5200 (Agilent Technology, Inc., Santa Clara, CA). RNA-sequencing
libraries were prepared using 200 ng of total RNA following a recently
developed 3’-based protocol, TM3’seq
(Pallares et al., 2020).
Libraries were amplified with 16 PCR cycles. All other procedures
followed the published protocol or manufacturer recommendations.
Libraries were combined in equimolar quantities and sequenced on one
lane of an Illumina NovaSeq S4 flow cell (Illumina Inc.) of 100 bp
single-end with an average of 2.17 million reads per sample mapping to
the transcriptome. Reads were mapped to the Macaca mulattareference assembly Mmul_10
(Warren et al., 2020) using
kallisto (v0.43.1) (Bray et
al., 2016). We chose the genome of Macaca mulatta , a closely
related cercopithecoid, as its richer annotation yielded a higher number
of mapped reads compared to the Theropithecus gelada genome
(Chiou et al., 2022).