Transcriptomics analysis of different porcine intestinal models (piglet jejunums, jejunum crypts, and organoids) and conventional cell line models upon infection with the transmissible gastroenteritis virus (TGEV)
Transcriptomics analysis of different porcine intestinal models (piglet jejunums, jejunum crypts, and organoids) and conventional cell line models upon infection with the transmissible gastroenteritis virus (TGEV)
ST cells were cultivated in EMEM (Lonza), 10% FBS, 1% Penicilinn/Streptomycin and 1% Glutamax (ThermoFisher). After two washes with EMEM but without FBS, ST cells (after 1 day of plating) were inoculated with TGEV at multiplicity of infection of 0.06. The cells were washed twice after 1 hour of inoculation and EMEM,1% Penicilinn/Streptomycin and 1% Glutamax was added. The corresponding controls were mock-infected with ST cells supernatant. The total RNA of jejunum crypts, 3D organoids, 2D organoids, piglet jejunums and ST cells were extracted with Trizol Reagent, according to the supplier's recommendations. After Turbo DNAse treatment and validation of RNA quality, 500ng of RNA were used to build the sequencing librairies. Libraries were generated with the Illumina Stranded mRNA Kit, according to the supplier's recommendations. Essentially, the mRNA were first enriched by the capture of polyA tails via oligoDT. After enzymatic fragmentation, the cDNA were synthetized and the protruding ends of the cDNA were converted into blunt ends by exonuclease/polymerase activities. The 3' ends were adenilated and the adaptators were ligated. The libraries were amplified by PCR reactions and sequenced.