title: Illumina HiSeq 2500 sequencing; RNA-seq for splenic gene expression in response to Newcastle disease virus challenge in Hy-Line Brown chickens
design: RNA-seq for splenic gene expression in response to Newcastle disease virus challenge in Hy-Line Brown chickens
library_construction_protocol: Spleen tissues were collected from each chicken at day 2 or day 6 post infection. At 21 days posthatch (dph), the chickens were divided randomly into two groups separated by different rooms. One group was treated with the LaSota lentogenic strain of NDV (200 ul of 108 EID50 for each chicken) through nasal and ocular inoculation routes (50 ul into each eye and nostril), while the control group was treated with 200ul phosphate-buffered saline (PBS) for each chicken through the same routes. RNA samples was extracted from spleen tissue using Ambion RNAqueous Total RNA Isolation Kit (Thermo Fisher Scientific) following the manufacturer's protocol. The transcriptome library of each sample was constructed with 0.5 µg total RNA using the Illumina TruSeq RNA sample preparation kit (Illumina Inc., San Diego, CA) following the low sample protocol in the TruSeq RNA sample preparation v2 guide (Part #15026495, March 2014).The proper length of cDNA in each library was guaranteed by running the DNA 1000 assay on the Agilent 2100 Bioanalyzer. Libraries from 2 and 6 dpi were independently constructed and sequenced at different times.