GSM4671000: WT_GF_229; Mus musculus; RNA-Seq
Adult microglia were harvested using density gradient separation. CD11b+CD45lo microglial cell populations were FACS-sorted and collected directly in cell lysis buffer. The SMARTer Ultra Low Input RNA Kit for Sequencing v4 (Clontech Laboratories, Inc., Mountain View, CA, USA) was used to generate first strand cDNA from 300 pg total-RNA. Double stranded cDNA was amplified by LD PCR (13 cycles) and purified via magnetic bead clean-up. Library preparation was carried out as described in the Illumina Nextera XT Sample Preparation Guide (Illumina, Inc., San Diego, CA, USA). 150 pg of input cDNA were tagmented (tagged and fragmented) by the Nextera XT transposome. The products were purified and amplified via a limited-cycle PCR program to generate multiplexed sequencing libraries. For the PCR step 1:5 dilutions of index 1 (i7) and index 2 (i5) primers were used. The libraries were quantified using the KAPA SYBR FAST ABI Prism Library Quantification Kit (Kapa Biosystems, Inc., Woburn, MA, USA). Equimolar amounts of each library were pooled, and the pools were used for cluster generation on the cBot with the Illumina TruSeq SR Cluster Kit v3.
GEO Accession
GSM4671000