RNA-Seq of bovine macrophages inoculated with Neospora caninum heat-killed Nc-Spain7 and Nc-Spain1H isolates
The poly(A)+ mRNA fraction was isolated from the total RNA and cDNA libraries were obtained following Illuminas recommendations. Briefly, poly(A)+ RNA was isolated on poly-T oligoattached magnetic beads and chemically fragmented prior to reverse transcription and cDNA generation. The cDNA fragments then went through an end repair process, the addition of a single A base to the 30 end and then ligation of the adapters. Finally, the products were purified and enriched using PCR to create the final indexed double-stranded cDNA library. The quality of the libraries was then analysed in a Bioanalyzer 2100, High Sensitivity assay; the quantity of the libraries was determined by real-time PCR in a LightCycler 480 (Roche Applied Science, Mannheim, Germany). Prior to cluster generation in cBot (Illumina, San Diego, USA), equimolar pooling of the libraries was performed. The pool of the cDNA libraries was sequenced by paired-end sequencing (100 bp x 2) in an Illumina HiSeq 2000 sequencer (Illumina, San Diego, CA, USA)
SUB5913685
CCM0_F_8B