GSM6857766: P7_10; Mus musculus; RNA-Seq
Single cell libraries were prepared according to the Smart-Seq2-protocol (22,23) with following modifications. Pre-amplification PCR cycles were increased to 23 to obtain sufficient amounts of cDNA for the sequencing analysis. Primer dimers were eliminated by two 0.8x Ampure-XP bead clean ups. 0.3-0.5 ng of pre-amplified cDNA were subjected to library preparation with the Nextera XT library preparation kit, Illumina, San Diego, CA, USA in 8 µl reaction volume. Barcoded libraries were pooled and sequenced the with a S1 flow cell and a 300-cycle kit on a NovaSeq Illumina platform to obtain 150-bp paired-end reads. Quality controls were performed using TapeStation with D5000 high sensitivity tapes (both from Agilent, Santa Clara, CA, USA). Quantification was performed using Qubit high sensitivity kit (Thermo Fisher Scientific) after PCR preamplification, and the PicoGreen assay (Thermo Fisher Scientific) after Nextera XT had been performed.