PT  - JOURNAL ARTICLE
AU  - Montagne, Janelle M.
AU  - Alice Zheng, Xuwen
AU  - Pinal-Fernandez, lago
AU  - Milisenda, Jose C.
AU  - Christopher-Stine, Lisa
AU  - Lloyd, Thomas E.
AU  - Mammen, Andrew L.
AU  - Larman, H. Benjamin
TI  - Ultra-Efficient Short Read Sequencing of Immune Receptor Repertoires
AID  - 10.1101/494062
DP  - 2018 Jan 01
TA  - bioRxiv
PG  - 494062
4099  - http://biorxiv.org/content/early/2018/12/24/494062.short
4100  - http://biorxiv.org/content/early/2018/12/24/494062.full
AB  - Immune receptor repertoire (IRR) sequencing is increasingly employed to characterize adaptive immune responses. However, current IRR sequencing methodologies are complex, expensive, or both, thereby limiting routine utilization. Here we present Framework Region 3 AmplifiKation sequencing (“FR3AK-seq”), a simplified multiplex PCR-based approach for the ultra-efficient analysis of IRRs. By using minimal primer sets that target a conserved region adjacent to the hypervariable VDJ sequence, undistorted amplicon can be analyzed via short read, single-end sequencing. We find that FR3AK-seq is sensitive and quantitative, with a per sample cost of ~50-fold below the current industry standard. Inference of V-allele usage from FR3AK-seq data is demonstrated using a novel algorithm: Inferring Sequences via Efficiency Projection and Primer Incorporation (“ISEPPI”). FR3AK-seq and ISEPPI were utilized to quickly and inexpensively characterize the T cell infiltrates of 146 muscle biopsies obtained from patients with idiopathic inflammatory myopathies (IIMs) and controls. A cluster of related T cell receptors were identified in samples from patients with sporadic inclusion body myositis, suggesting the presence of an unknown shared antigen. The ease and cost of FR3AK-seq analysis removes the current barriers to routine, large-scale IRR analyses.