Mk28 r

RNA-seq experiment was performed as previously reported (11 (link)). In vivo bone lesions were collected 4 weeks after inoculation. BICA samples were harvested and pooled 3 weeks after tumor seeding. All specimens were grinded and homogenized by Precellys 24 Homogenizer (Bertin instrument) with 2.8mm stainless steel beads in 2mL reinforced tubes (Bertin instruments, MK28R). Total RNA of cancer/bone mixture was extracted using the Direct-zol RNA miniprep kit (Zymo Research, R2051). The first cDNA strands were prepared by RevertAid First Strand cDNA Synthesis Kit (Thermo Scientific, K1622) with > 200 ng total RNA input. The second cDNA strand synthesis was performed with NEBNext mRNA Second Strand Synthesis Module (NEB, E6111L). Libraries were generated with the Illumina Nextera XT DNA Library Prep Kit (Illumina). Cluster generation was conducted with the Illumina Nextseq 500/550 high output v2 kit and sequenced on the Illumina Nextseq 500 equipment with the assistance of BCM Genomic and RNA Profiling Core. The RNA-seq data were analyzed with the assistance of Roswell Park Bioinformatics Resource.