Integrating Visium Spatial Transcriptomics

The raw ST reads were aligned to the same reference package as for the snRNA-seq dataset using SpaceRanger (version 1.2.2, 10x Genomics) software to generate spot barcode-to-gene feature matrix for downstream analysis. Individual ST object was created by Load10X_Spatial() function with Seurat (v3.2.2) package (146 (link)) and normalized with SCTransform() function (147 (link)). The merge() function was used to aggregate 16 slices into one ST object (Visi) containing images, and the Visi@assays$SCT@counts was pulled to create a separate ST object (VisiDot) by CreateSeuratObject() function to get aggregated spots without images. A processing pipeline similar to that of snRNA-seq was employed for the VisiDot object; specifically, Harmony() was applied over “IL01_uniqueID” variable to integrate 16 samples. To match the terminal MRI images, the orientation of the Visium images was corrected by transforming the image array stored at Visi@images$slice1@image, and the spot information stored at Visi@images$slice1$@coordinates was swapped accordingly (See GitHub post for detail). The microenvironment (ME) cluster annotated in the VisiDot object was transferred back to the Visi object for spatial visualization.

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Lin J.P., Brake A., Donadieu M., Lee A., Kawaguchi R., Sati P., Geschwind D.H., Jacobson S., Schafer D.P, & Reich D.S. (2023). A 4D transcriptomic map for the evolution of multiple sclerosis-like lesions in the marmoset brain. bioRxiv.

Publication Preprint 2023

SpaceRanger

Assays Gene Snrna Spatial visualization Spots

Corresponding Organization : National Institute of Neurological Disorders and Stroke

Other organizations : University of California, Los Angeles, Cedars-Sinai Medical Center, Neuropsychiatric Research Institute, University of Massachusetts Chan Medical School

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Variable analysis

independent variables

None explicitly mentioned

dependent variables

Spot barcode-to-gene feature matrix
Aggregated spots without images (VisiDot object)
Microenvironment (ME) cluster annotation

control variables

Reference package used for alignment
Seurat package (version 3.2.2) used for analysis
SCTransform() function used for normalization
Harmony() applied over 'IL01_uniqueID' variable to integrate samples
Orientation of Visium images corrected to match terminal MRI images

positive controls

None specified

negative controls

None specified

Annotations

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