RNA-Seq of Rattus norvegicus: 8 week-old mnedulla with area postrema
Briefly, sections from rat medulla at the level of the area postrema were used. Sections were cut in a pre-cooled cryostat at 10m thickness onto a Visium Spatial Gene Expression Slide (PN:2000233/SN:V12U07-300 from 10 x Genomics B.V., LeCarrefour, Dellaertweg 9D, 2316 WC Leiden, The Netherlands) with four 6.5mm x 6.5mm capture areas each containing ~5000 oligo-barcoded spots. Each spot covered a 55m area with a 100m center-to-center distance. Slides then underwent fixation and HE staining using the protocol CG000160_DemonstratedProtocol_MethanolFixationandHEStaining_RevC.pdf on the 10x Genomics webpage. The stained sections were immediatedly imaged on an IX83 Live Imaging Microscope (Olympus) at 10x magnification. The high resolution images were used as input for Space Ranger without further adjustment. Space Ranger is a set of analysis pipelines that process 10x Genomics Visium data with microscope images (https://www.10xgenomics.com/support/software/space-ranger/latest). All subsequent reactions were done using the protocol CG000239_VisiumSpatialGeneExpression_UserGuide_RevF.pdf on the 10x Genomics webpage. The resulting DNA libraries were then sequenced on the NextSeq2000 VH00748 platform and 230512_VH00748_35_AAAYJ7THV Flowcells according to the Visium Spatial Gene Expression Reagent Kits User Guide (10x Genomics protocol), Version F.  The 10x Genomics protocol recommended a sequencing depth/spot of at least 50,000 read pairs per tissue covered spot on the capture area.