Table 1 Comprehensive overview of NASQAR applications

Preprocessing

Merge counts files into a matrix format that is needed for many downstream analysis; also offers gene id/name conversion

base R packages

gene count files (Eg. output count files from htseq)

csv file containing a matrix of samples and corresponding gene counts

N/A

Preprocessing

Merge sample fpkm files into a matrix format; also offers gene id/name conversion

base R packages

sample gene fpkm files (Eg. sample fpkm files from Cufflinks)

csv file containing a matrix of samples and corresponding gene fpkm values

N/A

RNAseq (bulk)

Differential Gene Expression (DGE) analysis and optionally Surrogate Variable Analysis (SVA) for hidden batch effect detection

DESeq2, SVA

matrix of samples and gene counts (csv) and optionally a metadata table (csv)

VST matrix, Rlog matrix, DGE results table, Gene expression table, normalized counts matrix

Distance heatmap, PCA plots, MA plots, Gene expression boxplots, normalized counts heatmap, SVA plots

RNAseq (bulk)

Differential Gene Expression (DGE) analysis

edgeR, limma-voom

matrix of samples and gene counts (csv)

Gene expression table

PCA plots, Heatmaps, Scatter plots, Volcano plots, Gene expression boxplots

RNAseq (bulk)

Differential Gene Expression (DGE) analysis

DESeq2, edgeR, limma-voom

matrix of samples and gene counts (csv) and a metadata table (csv)

DGE results table, method comparison results

PCA plots, Dispersion plot, Volcano plot, Venn diagram method comparison

RNAseq (single-cell)

Guided single-cell RNA-seq data analysis and clustering.

Seurat, sctransform, dplyr

either 10X data files (mtx, tsv) OR matrix of cell/gene counts

PCA results, ICA results, DGE cluster markers table, Seurat R Object, R script

Violin plots, PCA plots, PCA heatmaps, Elbow Plot,UMAP, t-SNE

Metagonomics

Differential analysis of quantitative metagenomic data

DESeq2, circlize, ape, phytools, philentropy

a BIOM matrix file OR a counts file (csv)/taxonomy file (tsv). Optionally a phylogenetic tree newick file (nhx) and/or fastq files

Output files (biom, tsv, nhx) of Shaman workflow that can be used in downstream metagenomic analysis and visualization within Shaman

Bar plots, PCOA/PCA plots, Clustering dendograma, Rarefaction curves, Scatter plots, Heatmaps, Box plots, Diversity plots, Venn diagram

Gene Enrichment - Gene Set Enrichment Analysis

Gene Set Enrichment Analysis (GSEA) of GO-Terms and KEGG pathways

clusterProfiler, DOSE, Goplot, enrichplot, pathview

a table of differential gene expression DGE data (csv/tsv)

GO terms table, KEGG results table

Dot plots, Category netplot, GO induced graph, Pathview plot, ridge plot, pubmed trends

Gene Enrichment - Over Respresentation Analysis

Over-representation analysis of GO-Terms and KEGG pathways

clusterProfiler, DOSE, Goplot, enrichplot, pathview, wordcloud2

a table of differential gene expression DGE data (csv/tsv)

GO terms table, KEGG results table

Bar plots, Dot plots, Category netplot, GO induced graph, Pathview plot, word cloud