RNAseq of Lumenal-derived Teladorsagia circumcincta larvae-2-3529
RNA was extracted from three separate samples (biological replicates) of luminal or mucosal isolated T. circumcincta larvae using Trizol reagent (Life Technologies, UK) on dry ice. RNA quality was assessed on a spectrophotometer (Nanodrop, Thermo Scientific Ltd, UK) with OD260/280 ranging from 1.89-2.1. Samples of sufficient quality for subsequent analysis were obtained from three paired MD and LD populations from three individual sheep. RNA (5ľg of each sample) was processed for Illumina Solexa Hi-Seq sequencing following the manufacturerŐs protocol. Base calls were made using the Illumina CASAVA 1.8 pipeline. The quality of the reads was checked with FastQC v0.10.1 and sequences processed to remove primer and adapter sequences. The resulting sequences were then aligned to the T. circumcincta transcriptome using Samtools (version 0.1.18) allowing the assembled Illumina Hi-Seq sequences to be cross-referenced with the T. circumcincta transcriptome contig identities. The total number of reads for each of the six RNA sample sets, which mapped to each contig/isotig of the T. circumcincta transcriptome was then determined.