mixcr-analysis

Run mixcr analyze with the generic-amplicon preset on paired-end FASTQs, specifying species, --rna, and CDR3-based clonotype assembly. Then use mixcr exportClones with --chains to export TRA, TRB, TRD, and TRG tables for downstream R analysis.

Use qiagen-human-rna-tcr-umi-qiaseq only if barcodes are still in the R2 sequence. If FASTQs are already demultiplexed with barcodes in the header, switch to generic-amplicon to avoid over 90% barcode extraction failures.

MiXCR is intended here for bulk amplicon data, not single-cell. For 10x VDJ data, use a single-cell workflow such as the scRepertoire skill instead of this bulk processing pipeline.

Low alignment usually means the wrong species was specified, the reads contain non-TCR/BCR contamination, poor sequencing quality, or a mismatched preset. Check the align report with grep for 'Successfully aligned' and verify quality with FastQC.

Simpson clonality is 1 minus Shannon entropy normalized by log of clone count, ranging from 0 (perfectly diverse) to 1 (monoclonal). Healthy blood repertoires typically score 0.01-0.10, while tumor-infiltrating samples often exceed 0.10.

Empty TRD or TRG files are normal when samples are dominated by alpha-beta T cells, since gamma-delta chains are rare unless specifically enriched. Verify alignment succeeded first, then confirm the chain is biologically expected in your sample type.