Reproducibility and Reuse of Adaptive Immune Receptor Repertoire Data

Georgiou and colleagues discuss rapidly evolving methods for high-throughput sequencing of the antibody repertoire, and how the resulting data may be applied to answer basic and translational research questions.

IMGT/V-QUEST is the highly customized and integrated online IMGT(®) tool for the standardized analysis of the immunoglobulin (IG) or antibody and T cell receptor (TR) rearranged nucleotide sequences. The analysis of these antigen receptors represents a crucial challenge for the study of the adaptive immune response in normal and disease-related situations. The expressed IG and TR repertoires represent a potential of 10(12) IG and 10(12) TR per individual. This huge diversity results from mechanisms that occur at the DNA level during the IG and TR molecular synthesis. These mechanisms include the combinatorial rearrangements of the variable (V), diversity (D) and joining (J) genes, the N-diversity (deletion and addition at random of nucleotides during the V-(D)-J rearrangement) and, for IG, somatic hypermutations. IMGT/V-QUEST identifies the V, D, J genes and alleles by alignment with the germline IG and TR gene and allele sequences of the IMGT reference directory. The tool describes the V-REGION mutations and identifies the hot spot positions in the closest germline V gene. IMGT/V-QUEST integrates IMGT/JunctionAnalysis for a detailed analysis of the V-J and V-D-J junctions and IMGT/Automat for a complete annotation of the sequences and also provides IMGT Collier de Perles. IMGT/HighV-QUEST, the high-throughput version of IMGT/V-QUEST, implemented to answer the needs of deep sequencing data analysis from Next Generation Sequencing (NGS), allows the analysis of thousands of IG and TR sequences in a single run. IMGT/V-QUEST and IMGT/HighV-QUEST are available at the IMGT(®) Home page, http://www.imgt.org.

Driven by dramatic technological improvements, large-scale characterization of lymphocyte receptor repertoires via high-throughput sequencing is now feasible. Although promising, the high germline and somatic diversity, especially of B-cell immunoglobulin repertoires, presents challenges for analysis requiring the development of specialized computational pipelines. We developed the REpertoire Sequencing TOolkit (pRESTO) for processing reads from high-throughput lymphocyte receptor studies. pRESTO processes raw sequences to produce error-corrected, sorted and annotated sequence sets, along with a wealth of metrics at each step. The toolkit supports multiplexed primer pools, single- or paired-end reads and emerging technologies that use single-molecule identifiers. pRESTO has been tested on data generated from Roche and Illumina platforms. It has a built-in capacity to parallelize the work between available processors and is able to efficiently process millions of sequences generated by typical high-throughput projects.