GSM3985887: Sample9_chd-overexpressing_repeat3; Danio rerio; RNA-Seq
To prepare total RNA, the TRIzol reagent (Invitrogen) manufacturer's protocol was followed until aqueous phase recovery, then 6.25 μl Co-Precipitant Pink (Bioline) was added to 250 μl aqueous phase, followed by 375 μl 100% EtOH. After vortexing briefly, samples were transferred to RNeasy Mini kit (Qiagen) spin columns and centrifuged at 13600 rpm at 4°C for 1 min. Flow-through was discarded and columns were washed twice with RPE buffer (Qiagen). RNA was eluted in 50 μl H2O. Total RNA concentration was measured using a NanoDrop spectrophotometer (ThermoScientific). Libraries were prepared by LGC Genomics GmbH (Berlin, Germany) for Illumina NextSeq 500 V2 sequencing. For mRNA isolation and cDNA synthesis, the following kits were used: NEBNext Poly(A) mRNA Magnetic Isolation Module, NEBNext RNA First Strand Synthesis Module, and the NEBNext UltraII Non-Directional RNA Second Strand Synthesis Module. Resulting cDNAs were purified on MinElute Columns (Qiagen) and used to prepare sequecing libraries with the "Encore Rapid DR Multiplex System" (NuGen). Libraries were amplified (12 cycles) using standard Illumina primers.
GEO
GSM3985887