Comparison of Three Libraries for 10x Genomics Single Cell Immune TCR Repertoire Profiling.

10x Genomics single cell sequencing provides a comprehensive and scalable solution for cell characterization and gene expression profiling of hundreds to tens of thousands of cells. We tested three libraries: (1) 5 prime gene expression, (2) direct enrichment for TCR, and (3) post-cDNA-amplification enrichment for TCR, for 10x Genomics single cell immune TCR repertoire profiling on two Invariant natural killer T (iNKT) cells B240 and B241 with iNKT purity at 90% and 45% respectively. We sequenced about 1100 cells on B240, and 580 cells on B241. We identified the number of productive clonotypes at 165, 284, and 332 on B240 for three libraries respectively, where 47 clonotypes are overlapped among all three libraries. B241 has the number of productive clonotypes at 88, 152 and 148 for three libraries respectively, where 35 clonotypes are overlapped among all three libraries. We also performed the correlation between gene expression data and clonotypes with the data by library 3. Since each sample is a mixture of iNK T cells and classic NK T cells (non-invariant), PCA/tSNE plots of the 5' gene expression data shows two separate clusters for each sample, identifiable by TCR clonotype usage. One cluster contains unique or very few alpha/beta clones, while the other one contains diverse alpha/beta clones.Based upon our data, we noticed that the number of clonotypes identified is proportional to the number of cells sequenced regardless of libraries. Library 1 has low sensitivity in detecting clonotypes due to un-enrichment of TCR genes, and only beta chain was detected for clonotypes. The other two libraries are able to identify high and similar number of clonotypes. However, library 3 is able to associate gene expression pattern with TCR clonotype usage, which is a big advantage over other two libraries for 10x Genomics single cell immune TCR repertoire profiling.