title: Spatial Transcriptomics of Mus Musculus: PFF injected
design: Illumina Novaseq 6000 was used for sequencing with a read length of 27 for both reads with reverse sequence orientation in the readout group plate information. Plates were dried down and rehydrated in 10 L nuclease-free water, mixed and incubated at room temperature for 10 minutes. PCR was performed on samples as described in the nanoString GeoMx DSP Readout User Manual using 2 L PCR master mix, 4 L primer from the correct wells, and 4 L resuspended DSP aspirate. KAPA beads (KAPA Pure beads, Roche Cat# 07983298001) were warmed to room temperature for 30 minutes. Libraries were pooled and KAPA beads were added to each pool at a 1.2X ratio to the final pool volume. Two KAPA bead clean ups were performed and pooled libraries were eluted in 24 L elution buffer. Negative and positive control pools were eluted into 10 L elution buffer. Quantity of the pools were assessed using the QuantiFluor dsDNA System (Promega Corp.). Pools were diluted to 5 ng/L and quality and size are assessed using Agilent DNA High Sensitivity chip (Agilent Technologies, Inc.) on the Bioanalyzer. Sequencing was performed at 100 reads/m2. Paired end 50 base paired sequencing. Base calling was done by Illumina RTA3 and output was demultiplexed and converted to fastq format with bcl2fastq v1.9.0.