title: RNA-seq of mus musculus: alveolar macrophage
design: Typically, 1  g of total RNA was used with the TruSeq RNA library preparation kit (Illumina) in accordance with low-throughput protocol, except that SuperScript III reverse transcriptase (Invitrogen) was used to synthesize first strand cDNA. In the step of end repair the synthesized cDNA was subjected to end-repair and then was 3  adenylated. Adaptors were ligated to the ends of these 3  adenylated cDNA fragments. We used PCR to amplify the cDNA fragments with adaptors from previous step. PCR products are purified with the SPRI beads, and dissolved in EB solution. In the circularization step the double stranded PCR products were heat denatured and circularized by the splint oligo sequence. The single strand circle DNA (ssCir DNA) were formatted as the final library.