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Table 3 All methods implemented in IMDA (Fig. 1) for automated immune repertoire analysis

From: ImmunoDataAnalyzer: a bioinformatics pipeline for processing barcoded and UMI tagged immunological NGS data

Method Description Input (I)/Output (O)
MIGEC Read assignment by barcode (de-multiplexing) and consensus assembling based on UMIs using the open-source tool MIGEC. I: NGS files in compressed or non-compressed FASTQ file format O: assembled reads in FASTQ file format
MiXCR Execution of MiXCR commands for clonotype identification and quantification for receiving nucleotide and AA sequences of the CDR3 region and V(D)J gene segments. I: files in compressed or non-compressed FASTQ file format O: immune repertoire profiling measures (e.g., V(D)J gene segments, CDRs etc.) in text file format
ContaminationAnalysis Calculates shared UMIs and V(D)J hits of multiple samples for cross-sample contamination analysis. I: MiXCR output, MIGEC output or non-compressed FASTQ files containing the UMI sequence in the read ID O: cleaned FASTQ files
FACSCorrection Cell subset disambiguation of clonotypes from cells separated using FACS or magnetic sorting (e.g., CD4+ and CD8+) and elimination of clonotypes within a twofold change range. I: filename of pairs for analysis (pairs.txt) and MiXCR output O: cleaned files in MiXCR text file format
VDJtools Executes methods of the open-source tool VDJtools (convert and calculate diversity indices) for diversity stats visualization later-on. I: MiXCR output O: diversity indices for all samples
Bowtie Analyze undetermined reads from sequencing run and from MIGEC assignment using the open-source tool Bowtie2 for non-assignable read composition analysis. I: file path to the undetermined files in compressed or non-compressed FASTQ file format and to MIGEC undef-m output file, and path to Bowtie2 library O: mapping information is collected in SAM file format
EvaluateUndetermined If undetermined read analysis using Bowtie2 has been performed, evaluation and visualization of the results is done. I: the Bowtie2 output in SAM file format O: read counts for each category (see Undetermined Read Processing and Analysis)
ClonalityAnalysis Clonotype and CDR3 sequence length analysis and visualization is performed. I: MiXCR output O: CDR3 AA length distribution and clonotype counts
DiversityAnalysis Diversity curves are calculated and visualized as well as the diversity indices calculated using VDJtools. I: MiXCR output and VDJtools output O: diversity curves and diversity measures
OverlapAnalysis Shared clonotype analysis and visualization. I: MiXCR output O: shared clonotype overlaps (heatmap and LM plots)
SimilarityAnalysis Hierarchical clustering of all samples is performed and visualized. I: MiXCR output O: hierarchical clustering information
VDJAnalysis Calculation of V and J gene segment pairings and visualization using Chord diagrams. I: MiXCR output O: chord diagrams