GSM9082303: Male,4wks Saline, Sample 4; Mus musculus; RNA-Seq
Microglia were isolated using FACS. Total microglial RNA was extracted using the RNeasy Plus Micro Kit (Qiagen). RNA concentration was determined with the NanopDrop One spectrophotometer (NanoDrop) and RNA quality assessed with the Agilent Bioanalyzer 2100 (Agilent). 500 pg of total RNA was pre-amplified with the SMARTer Ultra Low Input kit v4 (Clontech) per manufacturer's recommendations. The quantity and quality of the subsequent cDNA was determined using the Qubit Fluorometer 3.0 (Life Technnologies) and the Agilent Bioanalyzer 2100 (Agilent). 150pg of cDNA was used to generate Illumina compatible sequencing libraries with the NexteraXT library preparation kit (Illumina) per manufacturer's protocols. The amplified libraries were hybridized to the Illumina flow cell and sequenced using the NovaSeq X sequencer (Illumina) with target depth of ~30 million of 150 nt paired-end reads per sample.