GSM2668622: DO2-3; Apostichopus japonicus; RNA-Seq
According to the instructions provided by the manufacturer of the RNase-free DNase (Qiagen Inc., Hilden, Germany), an RNeasy mini kit including DNase treatment was used to isolate total RNA from nine samples (three replicates in each group) of the respiratory tree from the three different groups (DO2, DO4, and DO8). The RNA samples from three individuals per replicate were pooled in equal amounts for library construction. Libraries were prepared from the nine RNA pools by Beijing Genomics Institute, Shenzhen, China (BGI-Shenzhen). Strict quality control (QC) was applied in each experimental step to obtain highly reliable sequencing data. Magnetic beads with poly-T-oligos attached were used to isolate poly(A) mRNA. The target RNA was then purified. The target RNA was fragmented into short sequences, reverse-transcribed into single-strand cDNA using the N6 random primer (Promega, Madison, WI, USA), and synthesized into double-stranded cDNA (dscDNA) with DNA polymerase I and RNase H (Promega, Madison, WI, USA). Next, the end of the dscDNA was repaired and ligated with a sequencing adapter. The ligation product was amplified using specific primers. The PCR product was denatured by heating and the single strand DNA was cyclized using splint oligo and DNA ligase. Finally, high-throughput sequencing was performed with the prepared cDNA libraries using the BGISEQ-500 RS platform at BGI-Shenzhen.
GEO Accession
GSM2668622