title: Nanopore direct RNA-seq of chicken female spleen
design: The direct RNA libraries were prepared using the Direct RNA Sequencing Kit SQK- RNA002 (Oxford Nanopore Technologies) according to the manufacturers protocol. Briefly, the poly-A RNAs were enriched using the NEBNext Poly(A) mRNA Magnetic Isolation Module (CAT#E7490, NEB, USA) with NEBNext Magnetic Oligo (dT)25 Beads. Next,100500ng of the poly-A enriched RNAs were ligated to the reverse transcriptase adaptor using T4 DNA ligase, followed by reverse transcription. The reverse-transcribed RNAs were ligated to the sequencing adaptor and were purified using Agencourt RNAClean XP beads (Beckman Coulter). Finally, two libraries (for brain and spleen) were constructed and sequenced on two different R9.4.1 FlowCells using the PromethION sequencer (ONT, UK) at Grandomics Biosciences (Wuhan, China).