Data upload & pipeline configuration
Upload your 10x Genomics zip files and mitochondrial gene list. Each zip should contain barcodes.tsv.gz, features.tsv.gz, and matrix.mtx.gz at its root level.
Mitochondrial gene list

CSV with a column named 'GeneName'

10x replicate files (zip)
Replicate 1 required
Replicate 2 optional
Replicate 3 optional
QC thresholds
Cells outside these ranges are removed. Violin plots on the QC tab will show the effect of your choices. Start with defaults and adjust if needed.
Clustering parameters
Resolution guide: 0.3–0.5 = fewer, broader clusters (good starting point for most midgut datasets). 0.8–1.2 = more granular subclusters.
Integration & correction method
CCA Integration recommended
Seurat anchor-based. Best for 2–3 replicates sharing cell types. Produces a shared 'integrated' assay.
Harmony faster
Corrects PCA embeddings iteratively. Better for large datasets (>50k cells) or many batches.
Both methods produce a UMAP. You can compare integration quality on the UMAP tab after the run.
Optional analyses
Configuration summary


Typical run times:
10,000 cells → ~8 min
30,000 cells → ~25 min
50,000 cells → ~45 min
Do not close the browser tab while running.
Pipeline progress
Dataset statistics
Total cells
post-QC + doublet removal
Clusters
Replicates
uploaded
Markers found
across all clusters
Pre- vs post-QC violin plots
These violin plots show the distribution of nFeature_RNA, nCount_RNA, and percent.mt before and after applying your QC thresholds. Ideal post-QC distributions are tighter and centred.
Download QC plot
UMAP coloured by sample
Good integration: replicate colours should be mixed throughout the UMAP, not forming separate islands.
Download
UMAP coloured by cluster
Each number is a Seurat cluster. Use the Marker Genes tab to identify cell types in each cluster.
Download
Cluster cell count
Replicate contribution per cluster
Gene expression visualisation


Gene names are case-sensitive. Check your species notation (e.g. Drosophila uses lowercase for most genes).
Feature plot (expression on UMAP)
Download
Violin plot (expression per cluster)
Download
Monocle3 pseudotime trajectory
What is pseudotime? Cells are ordered along a trajectory from a root population (e.g. stem cells) towards differentiated states. Colour intensity = how far a cell is along the trajectory. Earlier = darker.
Pseudotime on UMAP
Download
Top pseudotime-associated genes
Genes with Moran's I > 0.1 and q < 0.05 in the graph test are shown. These change significantly along the trajectory.
Download full table

Cluster marker genes
Markers are genes significantly upregulated in one cluster vs all others (Wilcoxon test, min.pct=0.25, log2FC>0.25). Use avg_log2FC and pct.1 to prioritise candidates.
Download this cluster CSV
Download ALL clusters CSV

Top 5 markers — dot plot
Seurat object

The final integrated Seurat object with all assays, reductions, and cluster identities. Load in R with readRDS().

Download Seurat object (.rds)
All marker genes

Complete FindAllMarkers() table — all clusters, all genes, sorted by cluster and log2FC.

Download all markers (.csv)
Analysis report checklist