title: GSM4724946: CD11b+  size-matched input rep2; Mus musculus; OTHER
library_construction_protocol: CD11b+ cells were obtained by harvesting the splenic tumors from the WT/MLL-Af4 or I3/MLL-Af4 mice, lysing in red blood cell lysis buffer, and staining with a CD11b biotinylated antibody for positive selection by MACS (Miltenyi). RNA from both cell types was collected using QIAzol Lysis Reagent and purified using the miRNeasy Mini Kit (217004; Qiagen). The Agilent 2100 Bioanalyzer using RNA pico chip was utilized to quality check total RNA. The Nugen Universal plus mRNA-Seq Kit was used to generate strand-specific RNA-seq libraries. Samples were subjected to poly(A) RNA selection, RNA fragmentation and double-stranded cDNA generation using a mixture of random and oligo(dT) priming. The cDNA samples were then subjected to end repair to generate blunt ends, adapter ligation, strand selection, and PCR amplification to generate the final library. To mulitplex samples in one sequencing lane, different index adapters were used. Single-end, strand-specific RNA sequencing was performed on the Illumina HiSeq3000 system for the Lin- and CD11b+ samples, resulting in 15-20 million reads per sample, at the the UCLA Technology Center for Genomics & Bioinformatics. Nugen Universal plus mRNA-Seq Kit (RNA-seq) and Eclipse BioInnovations Inc eCLIP kit (eCLIP)