This is a simultaneous analysis of genes and Transposable elements (TEs) from 10X data, using the lovain clustering. We performed the analysis using “Alevin” as a tool integrated with the salmon software that uses EM algorithms for the analysis of 3’ tagged-end single-cell sequencing data.
1. The code named Preparing_files_for_Alevin_bash.sh is step-wise codes where we indexed the concatenated genes and TEs transcriptome and genome reference file using salmon.
2. we run Alevin on all the 10X generated samples at once using Running_Alevin.sh
3. Next file Imorting_Merging_files_fromAlevin_output.r is where we import and merge all datasets in one seurat object. We also calculate the percentage of mitochondrial or ribosomal genes, and the statistics of numbers of genes, UMIs in single cells
4. Now we remove the doublets, and annotate the cells from merged but not integrated objects using the codes in Doublet_removed_Annotating_cells_from_merged_object.r
DATA ANALYSIS -
Once the datasets were merged or even as individual files, we performed the integration, normalization, dimension reduction, visualization, and trajectory analysis of the single cellular clusters. Also, TE loci graphs were generated using the normalized data. All of the relevant codes are present in the file Data_integration_Dimension_reduction_visualization.r
TE family level analysis
This is followup of previous code where we analyze the data for TEs at family level. Also by removing all the TEs and comparing with genes only analysis. Code is given in TE_analysis_at_family_level_and_only_genes_analysis.r