title: GSM5047302: KL rep1; Mus musculus; RNA-Seq
library_construction_protocol: Single-cell suspension were isolated from tumor-bearing KL mice using micro-dissection after 10 weeks of Ad-cre induction. And cells were prepared as described above. Single-cell RNA-seq libraries were prepared with Chromium Single cell 3'Reagent v3.0 Kits according to the manufacturer's protocol. Single-cell suspensions were loaded on the Chromium Single Cell Controller Instrument (10×Genomics) to generate single cell gel beads in emulsions (GEMs). Briefly, 105 single cells were suspended in calcium- and magnesium-free PBS containing 0.04% weight/volume BSA. About 15000-20000 cells were added to each channel with a targeted cell recovery estimate of 5000 cells (4400 and 2780 for 2 KL mice, respectively). After generation of GEMs, reverse transcription reactions were engaged barcoded full-length cDNA followed by the disruption of emulsions using the recovery agent and cDNA clean up with DynaBeads Myone Silane Beads (Thermo Fisher Scientific). cDNA was then amplified by PCR with appropriate cycles which depend on the recovery cells. Subsequently, the amplified cDNA was fragmented, end-repaired, A-tailed, index adaptor ligated and library amplification. Then these libraries were sequenced on the Illumina sequencing platform (HiSeq X Ten), and 151 bp paired-end reads were generated. single cell RNA libraries were prepared for sequencing using standard Illumina protocols scRNA-Seq