NHS-IL12-treated cells, Replicate 2
TC-1/a9 cells (5_10^4) were implanted subcutaneously into the right flank of female C57Bl/6 mice. Mice were randomized and treatment was initiated when tumors reached 50-80 mm^3.  Tumors were homogenized using the Tissue Ruptor II (Qiagen) system with disposable probes.  Tumor RNA was isolated with RNeasy kit (Qiagen) in conjunction with RNase-free DNase digestion kit (Qiagen).  Tumor bulk RNA was sequenced by Novogene UC Davis Sequencing Center (Novogene Corporation, Ltd). Briefly, RNA samples were assessed for integrity and quantified using the Nano 6000 Assay Kit for the Bioanalyzer 2100 system (Agilent Technologies).  The NEBNext Ultra RNA Library Prep Kit for Illumina was utilized for generating sequencing libraries.  First and second strand cDNA synthesis were performed, and adaptors ligated to the adenylated 3 ends of DNA.  Standardization of fragment sizes was carried out by selecting fragments of 150-200 bp for PCR.  PCR was performed and final library quality was assessed utilizing the Agilent Bioanalyzer 2100 system.  Clustering was carried out utilizing the cBot Cluster Generation System using PE Cluster Kit cBot-HS (Illumina) according to manufacturers instructions.  Library preparations were sequenced, and paired-end reads were generated on an Illumina platform.