RNA-Seq of human vaginal epithelial cells
Total RNA was assessed for quality using an Agilent Tapestation 4200, and samples with an RNA Integrity Number (RIN) greater than 8.0 were used to generate RNA sequencing libraries using the Illumina Stranded mRNA Prep (Illumina, San Diego, CA). Samples were processed following manufacturers instructions, starting with 500 ng of total RNA. Oligo dT beads were used for PolyA selection and random hexamer primers were used for reverse transcription and amplification. Resulting libraries were multiplexed and sequenced with 100 basepair (bp) Paired End reads (PE100) to a depth of approximately 25 million reads per sample on an Illumina NovaSeq 6000. Samples were demultiplexed using bcl2fastq Conversion Software (Illumina, San Diego, CA).