RNA-Seq of mus musculus
Cultured C2C12 cells were collected and seeded in 6-well plates (2  105 cells/well). The plates were incubated at 37C and 5% CO2 for 24 h. E-rhBMP-2 and C-rhBMP-2 solutions were prepared at a concentration of 100 ng/mL by diluting with DMEM containing 2% BS. After treating each well of the 6-well plate with the prepared solutions, the plate was incubated at 37C and 5% CO2. The cells were collected after 3, 6, 12, and 24 h of incubation and mRNA was extracted as described in the qRT-PCR analysis section. The acquired mRNA was purified using the Dynabeads mRNA Purification Kit (Invitrogen) to deplete rRNA and enrich poly(A)+ RNA using oligo d(T). Enriched mRNA was used for library construction using the MGIEasy RNA Directional Library Prep Kit (MGI, Shenzhen, China), according to the manufacturers instructions. The directional RNA libraries were then sequenced using the DNBSEQ-T7 sequencing instrument (BGI Genomics, China), according to the manufacturers instructions, yielding 150 bp paired-end reads.