GCTB_single-cell RNA sequencing (scRNA-seq) reference annotation

This notebook performs single-cell RNA sequencing (scRNA-seq) reference annotation analysis using 10X Chromium datasets derived from Giant Cell Tumor of Bone (GCTB) samples. Workflow overview:- Loading 10X Chromium datasets using Scanpy- Quality control (QC) and filtering of low-quality cells- Mitochondrial content calculation and visualization- Generation of filtered AnnData (.h5ad) files for each sample- Batch integration and metadata annotation- Dimensionality reduction (PCA, UMAP)- Leiden clustering analysis- Marker gene identification using Wilcoxon rank-sum tests- Construction of an integrated reference single-cell atlas Samples corresponding to `sc_adata9` and `sc_adata10` used modified filtering criteria, likely because they were post-treatment samples. Standard filtering criteria for most samples:- `min_counts = 1000` Modified filtering criteria for `sc_adata9` and `sc_adata10`:- `min_counts = 500` Common QC/filtering criteria applied across samples:- `min_genes = 100`- `min_cells = 30`- `max_counts = 80000`- `pct_counts_MT < 2` Additionally:- `sc_adata4` was excluded from downstream integration because it was labeled as a poor-quality sample. Main libraries used:- scanpy- scvi- numpy- pandas- seaborn- matplotlib- huetracer Input paths:- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/SK286_01/count/sample_filtered_feature_bc_matrix- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/SK286_04/count/sample_filtered_feature_bc_matrix- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/GCTB_06/count/sample_filtered_feature_bc_matrix- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/GCTB_07/count/sample_filtered_feature_bc_matrix- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/GCTB_08/count/sample_filtered_feature_bc_matrix- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/GCTB_09/count/sample_filtered_feature_bc_matrix- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/GCTB_11/count/sample_filtered_feature_bc_matrix- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/GCTB_12/count/sample_filtered_feature_bc_matrix- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/SK286_11/count/sample_filtered_feature_bc_matrix- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/SK304_04/count/sample_filtered_feature_bc_matrix Intermediate file paths:- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/output/reference_output/sc_adata1.h5ad- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/output/reference_output/sc_adata2.h5ad- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/output/reference_output/sc_adata3.h5ad- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/output/reference_output/sc_adata4.h5ad- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/output/reference_output/sc_adata5.h5ad- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/output/reference_output/sc_adata6.h5ad- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/output/reference_output/sc_adata7.h5ad- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/output/reference_output/sc_adata8.h5ad- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/output/reference_output/sc_adata9.h5ad- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/output/reference_output/sc_adata10.h5ad Final output paths:- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/output/reference_output/GCTB_reference_single_cell.h5ad- /mnt/QNAP1/data/giant_cell_tumor_sue/chromium/output/reference_output/GCTB_reference_single_cell.h5ad.sha256