Question: Is it necessary to sequence the Gene Expression library in a Barcode Enabled Antigen Mapping (BEAM) experiment?
Answer: Yes, a Gene Expression (GEX) library is required to analyze Barcode Enabled Antigen Capture (BEAM)/Antigen Capture data in Cell Ranger. The cellranger multi pipeline runs into an error when a GEX library is not provided.
If gene expression profiles are not of particular interest, you may perform shallow sequencing. A suggested starting point for sequencing the GEX library is 5,000 reads per cell. However, the minimum GEX sequencing depth needed for accurate cell calling may vary based on the sample type and data quality. Please read the 10x Genomics Technical Note CG000148 to learn about how sequencing depth and cell number influence the detection of major cell types in peripheral blood mononuclear cells (PBMCs).
Some additional advantages to including a GEX library:
- When library or sample quality is poor, including a GEX library (even with shallow sequencing) makes it easier to troubleshoot any issues that may arise with BEAM data.
- The stringent cell calling algorithm implemented by the cellranger vdj pipeline tends to exclude cells with low-quality contigs. Therefore, T or B cells with incomplete rearrangements (seen in certain leukemias and/or developmental stages) are often excluded from the call set. These barcodes are retained in the GEX library.
- In general, the presence of a GEX library improves the characterization of the cells and, therefore, the robustness of data evaluation.
Products: Single Cell Immune Profiling, Barcode Enabled Antigen Mapping (BEAM)