Single-cell RNA-seq data analysis

After library construction and sequencing, raw sequencing data were de-multiplexed and mapped to the mouse reference genome (mm10) using the CellRanger toolkit (10X Genomics, version 4.0.0). Gene expression matrices were then generated from Foxc2^fl/fl control and NC-Foxc2^-/- mice. The matrix files were then used for data processing and downstream analysis using the BIOMEX browser-based software platform and its incorporated packages developed in R (76 (link)). Quality control and data pretreatment was performed in BIOMEX with the following manually set parameters: (i) genes with a row average of <0.005 were excluded for downstream analysis and (ii) cells in which over 8% of unique molecular identifiers were derived from the mitochondrial genome were considered as dead cells and removed from downstream analysis. The data were then normalized in BIOMEX using similar methodology to the NormalizeData function as implemented in the Seurat package (77 (link)).

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Ujiie N., Norden P.R., Fang R., Beckmann L., Cai Z., Kweon J., Liu T., Tan C., Kuhn M.S., Stamer W.D., Aoto K., Quaggin S.E., Zhang H.F, & Kume T. (2023). Differential roles of FOXC2 in the trabecular meshwork and Schlemm’s canal in glaucomatous pathology. Life Science Alliance, 6(9), e202201721.

Publication 2023

CellRanger toolkit

Cells Gene expression Genes Library Mice Mitochondrial genome

Corresponding Organization : Northwestern University

Other organizations : Duke University, Hamamatsu University School of Medicine

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Variable analysis

independent variables

Genotype (Foxc2 fl/fl control vs. NC-Foxc2-/- mice)

dependent variables

Gene expression profiles

control variables

Mouse reference genome (mm10)
CellRanger toolkit (10X Genomics, version 4.0.0) for data processing
BIOMEX browser-based software platform and its incorporated packages developed in R for data analysis
Genes with a row average of <0.005 were excluded for downstream analysis
Cells in which over 8% of unique molecular identifiers were derived from the mitochondrial genome were considered as dead cells and removed from downstream analysis
Data normalization methodology similar to the NormalizeData function as implemented in the Seurat package

positive controls

Not explicitly mentioned

negative controls

Not explicitly mentioned

Annotations

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