BmN4_RNA
The RNA-seq library was constructed using the mRNA-seq Sample Preparation Kit according to the manufacturer's instructions (Illumina). Briefly, RNA was subjected to poly(A) selection using Sera-Mag Magnetic Oligo-dT Beads. Poly(A+) RNA was partially degraded by incubating in Fragmentation Buffer at 94 degree_C for 5 min. First-strand cDNA was synthesized using random primer and SuperScript II (Invitrogen), and second strand cDNA was synthesized using RNaseH and DNA pol I (Illumina). Double-stranded cDNA was size fractionated by 6% PAGE and cDNAs of 250micro300 bp were recovered. Illumina GA sequencing adaptors were ligated to cDNA ends. cDNAs were amplified by 15 cycles of PCR reactions using Phusion DNA Polymerase (Finnzymes). Thirty-six-base-pair single-end-read RNA-seq tags were generated using an Illumina GA sequencer according to the standard protocol.