title: MiSeq sequencing of rearranged heavy chain from Oncorhynchus mykiss: Spleen fish 4
design: Total RNA was extracted from gills, spleen and gut using a combination of TRI-Reagent and RNeasy Mini Kit (Qiagen).  To obtain cDNA from each sample, 2.5 g of RNA were reverse transcribed using RevertAid Reverse Transcriptase (Thermo Fisher Scientific) and oligo(dT). cDNA samples from spleen, gills and gut obtained from four representative fish were individually amplified using combinations of forward primers specific for each subgroup of IGHV genes and reverse primers specific for IGHM, IGHD or IGHT genes (C ,C? or C?). A library was constructed per individual tissues with the TruSeq DNA PCR-Free Library Prep Kit (Illumina). Libraries were pooled together and paired-end sequencing was performed on an Illumina MiSeq with a MiSeq Reagent Kit v3 (2 x 300 cycles) cartridge (Illumina).
library_name: Spleen_4