title: GSM5701748: Vehicle treated BMDM; Mus musculus; RNA-Seq
library_construction_protocol: Macrophages were harvested with Tryple (10 min, RT) before removing dead cells with the Easy Sep Dead Cell Removal Kit. Cells were washed twice with PBS/2% FBS and filtered over a 40 um filter. Libraries were prepared according to Chromium's instructions.Briefly, samples and reagents were partitioned into droplets. Within each droplet, the cell is lysed and incubated with reagents (oligos and master mix) and a reverse transcriptase reaction performed to generate full length barcoded cDNA from polyA-tailed mRNA transcripts. The droplets are then broekn and the cDNA molecules allowed to pool. This step is followed by bulk PCR-amplificaiton and enzymatic fragmention follow. Size selection is then used to optimize the insert size of the double-stranded cDNA prior to library construction. During library construction, Read 2 is added by Adapter ligation. Illumina P5 and P7 sequences and sample index sequences are added during the Sample Index PCR. In addition to these sequences required for Illlumina bridge amplification and sequencing, each fragment contains the 10x barcode, a unique molecular identifier added during the reverse transcriptase step and the cDNA insert seuqence used in the data analysis.