RNA-seq of Ctenopharyngodon idella cell line GCSB1441
Fresh-frozen cell lines were lysed in 1 ml TRIzol LS reagent (Life Technologies) and shaken vigorously for 10 min at room temperature. After adding 200 l chloroform (Carl Roth GmbH & Co. KG) and centrifuging at 13,000  g, 10 min, 4C, 200 l of the upper aqueous phase was used for RNA extraction. Total RNA was extracted using the Agencourt RNAdvance Tissue Kit (Beckman Coulter) with a KingFisher Flex Purification System (Thermo Fisher Scientific) according to the manufacturers instructions. DNase I digestion (Qiagen) was carried out prior to the final elution to remove genomic DNA residues. The quantity of total RNA was determined using the NanoDrop Lite Spectrophotometer (Thermo Fisher Scientific). Poly(A) RNA was isolated using the Dynabeads mRNA DIRECT Purification Kit (Invitrogen) following the manufacturers instructions. Fragmentation and strand-specific construction of whole transcriptome libraries were performed using the Collibri Stranded RNA Library Prep Kit for Illumina Systems (Invitrogen). The quality and integrity of the RNA and the final library were verified using the Agilent TapeStation 4150 (Agilent Technologies) with appropriate chips and reagents.