NextSeq 550 paired end sequencing; Role of Macrophage migration inhibitory factor (MIF) in pancreatic stellate cells (PSCs)
Role of Macrophage migration inhibitory factor (MIF) in pancreatic stellate cells (PSCs)
Mouse pancreatic stellate cells (Mif+/+ or Mif-/- mPSCs) were isolated from pancreas tissue in laminar hood and subjected to collagenase digestion and gradient centrifugation method. Mice were sacrifice by CO 2 asphyxiation and pancreatic tissues were collected in PBS containing 1% penicillin/streptomycin under aseptic conditions. Mif -/- - and Mif +/+ -mPSCs were grown in SteCM with growth factors and 2% serum. Cells were harvested in Trizol reagent and total RNA was extracted from Mif+/+ or Mif-/-mPSCs (three biological replicates) using Direct-zol™ RNA Miniprep following manufacturer's instructions (Zymo Research). After assessing the quality of RNA, 850 ng of total RNA was used for library preparation using unstranded TruSeq RNA Library Prep Kit v2 (Illumina) adhering to manufacturer's instructions.