An open protocol for modeling T Cell Clonotype repertoires using TCRβ CDR3 sequences

Abstract

Abstract T cell receptor repertoires can be profiled using next generation sequencing (NGS) to measure and monitor adaptive dynamical changes in response to disease and other perturbations. Genomic DNA-based bulk sequencing is cost-effective but necessitates multiplex target amplification using multiple primer pairs with highly variable amplification efficiencies. Here, we utilize an equimolar primer mixture and propose a single statistical normalization step that efficiently corrects for amplification bias post sequencing. Using samples analyzed by both our open protocol and a commercial solution, we show high concordance between bulk clonality metrics. This approach is an inexpensive and open-source alternative to commercial solutions.

Keywords

• Multiplex PCR
• Amplification bias
• Clonotype counts
• TCR sequencing
• Normalization
• Negative binomial