mRNA-Seq of canis lupus familiaris: whole blood
Peripheral blood sample collected in Qiagen RNAprotect Animal Blood Tube (Qiagen) of one critically ill dog with septic peritonitis secondary to foreign body ileus. Total RNA was extracted using RNeasy Protect Animal Blood Kit (Qiagen) according to the manufacturers protocol. The RNA Integrity Numbers and RNA concentration were determined by RNA ScreenTape system with 2200 Tapestation (Agilent Technologies, Santa Clara, CA, USA) and RNA HS Assay Kit with Qubit 3.0 Fluorometer (Thermo Fisher Scientific, Waltham, MA, USA), respectively. For mRNA-Seq library construction, KAPA Stranded mRNA-Seq Kit for Illumina (Roche) was applied according to the manufacturers protocol. The quality and quantity of the library was determined by using High Sensitivity DNA 1000 ScreenTape system with 2200 Tapestation (Agilent) and dsDNA HS Assay Kit with Qubit 3.0 Fluorometer (Thermo Fisher), respectively. Pooled libraries were diluted to 1.8 pM for 2x81 bp paired-end sequencing with 150-cycle High Output v2.5 Kit on the NextSeq 550 Sequencing System (Illumina) according to the manufacturers protocol.