title: GSM2454142: RNA NMP T+ S+; Mus musculus; RNA-Seq
library_construction_protocol: Total RNA from approximately 2,500 cells was isolated using the RNeasy Micro kit (Qiagen) and genomic DNA was digested on column with the addition of an extra 1ul of RNase-free DNase I (Roche) to ensure complete digestion. RNA-Seq libraries with the RNA equivalent of approximately 1,000 cells were generated using the TotalScript RNA-Seq Kit (Epicentre) with random hexamers according to manufacturer’s instructions with slight modifications. Briefly, all reagents during the cDNA synthesis steps were scaled down in order to achieve a total volume of 39µl, allowing all of the material to be used in the subsequent tagmentation procedure. To further improve strand-specificity of the libraries, Actinomycin D was added to the first strand cDNA synthesis reaction at a 5ng/µl final concentration and the libraries were finally amplified using 14 cycles.