title: RNA-Seq of VSV Field Isolate 22-25
design: The TruSeq Stranded mRNA Sample Preparation Kit (Illumina, San Diego, CA) was used for next generation sequencing library construction per manufacturers protocols with modifications. Briefly, RNA was fragmented for 5 minutes without polyA selection. First-strand cDNA synthesis was performed with random hexamer priming followed by second-strand cDNA synthesis using dUTP incorporation for strand marking. cDNA was purified using the Qiagen RNeasy MinElute kit before proceeding with library construction. End repair and 3` adenylation was then performed on the double stranded cDNA. Illumina adaptors were ligated to both ends of the cDNA, purified by AMPure bead size selection, and amplified for 15 cycles of PCR with primers specific to the adaptor sequences to generate the final library of cDNA amplicons of approximately 200-500bp in size.