Update README.md

README.md

@@ -5,6 +5,24 @@ Amodio G, Giacomini G, Boeri L, et al. **T cell exhaustion and senescence signat
 Single-cell (from 10X Genomics) analysis of CD3+ T cells purified from the peripheral blood of men diagnosed with oligo-astheno-teratozoospermia (OAT, n=4), idiopathic non-obstructive azoospermia (iNOA, n=6), and a control group (FER, n=5).
 
  
+
+**scRNAseq** analysis was performed using a standard [Seurat](https://satijalab.org/seurat/) pipeline that includes the following steps starting from a minimal object after loading of 10X data to markers identification:
+
+   - Preprocessing and cell filtering
+      - Each sample was pre-processed and cells with mitochondrial RNA percentages higher than 10 and a number of features <1200 or >6000, were filtered out.   
+   - Normalization 
+      - Default seurat settings [vignette]
+   - Scaling: 
+      - Data was regressed out by passing UMI count, the percentage of mitochondrial genes, the difference between the cell cycle phases scores.
+   - Dimensionality reduction and Harmony batch removal:
+      - A principal component analysis (PCA) with 100 principal components (PCs) was performed and a UMAP-representation as well as clusters were computed on the top 55 components (orig.ident as batch variable)
+   - Clustering
+   - Markers identification
+      - Cluster annotation: 
+      - Intra-cluster comparisons
+
+  
+
 ### Directories and Files ###
 - **Script**: R scripts used for the analyses
   - `1_PreProcessing_Data.R`: Full object creation