This repository contains the bash command lines for processing the raw data and R command lines for the analyses that were generated for the Whelan Lab paper <>. Raw data and matrices are available in GEO <>. Analyses were done chiefly using Seurat(v3) and Monocle(v2). Additional libraries are specified throughout.
There are 5 walkthroughs:
- PreProcess_and_Alignment (Bash lines for read deconvolution, genome alignment, feature counting, and matrix generation for downstream analyses)
- Initial_clustering_Seurat (Initial pre-processing and scRNA-Seq QC using Seurat, as well as some early visualizations of the data)
- Monocle_PT_Epi (Pseudotime command lines using Monocle for all epithelial cells, along with BEAM)
Note that functions defined in Monocle_PT_Epi are used in Monocle_PT_Basal
- Monocle_PT_Basal (Pseudotime for just basal cells, and BEAMs associated with it)
- Downstream_Seurat (Additional Seurat analyses for data validation, miscellaneous differential expression tests, and marker validations)
Major requirements:
Bash:
UMI-Tools (1.0.0): https://umi-tools.readthedocs.io/en/latest/
STAR (2.7): https://github.com/alexdobin/STAR/blob/master/doc/STARmanual.pdf
Subread (2.0): http://subread.sourceforge.net/
R:
Seurat (3.1.1): https://satijalab.org/seurat/
Monocle (2.16.0): http://cole-trapnell-lab.github.io/monocle-release/docs_mobile/