Unknown

Dataset Information

0

A facile technology for the high-throughput sequencing of the paired VH:VL and TCR?:TCR? repertoires.


ABSTRACT: Natively paired sequencing (NPS) of B cell receptors [variable heavy (VH) and light (VL)] and T cell receptors (TCRb and TCRa) is essential for the understanding of adaptive immunity in health and disease. Despite many recent technical advances, determining the VH:VL or TCRb:a repertoire with high accuracy and throughput remains challenging. We discovered that the recently engineered xenopolymerase, RTX, is exceptionally resistant to cell lysate inhibition in single-cell emulsion droplets. We capitalized on the characteristics of this enzyme to develop a simple, rapid, and inexpensive in-droplet overlap extension reverse transcription polymerase chain reaction method for NPS not requiring microfluidics or other specialized equipment. Using this technique, we obtained high yields (5000 to >20,000 per sample) of paired VH:VL or TCRb:a clonotypes at low cost. As a demonstration, we performed NPS on peripheral blood plasmablasts and T follicular helper cells following seasonal influenza vaccination and discovered high-affinity influenza-specific antibodies and TCRb:a.

SUBMITTER: Tanno H 

PROVIDER: S-EPMC7176429 | biostudies-literature | 2020 Apr

REPOSITORIES: biostudies-literature

altmetric image

Publications


Natively paired sequencing (NPS) of B cell receptors [variable heavy (VH) and light (VL)] and T cell receptors (TCRb and TCRa) is essential for the understanding of adaptive immunity in health and disease. Despite many recent technical advances, determining the VH:VL or TCRb:a repertoire with high accuracy and throughput remains challenging. We discovered that the recently engineered xenopolymerase, RTX, is exceptionally resistant to cell lysate inhibition in single-cell emulsion droplets. We ca  ...[more]

Similar Datasets

| S-EPMC5801115 | biostudies-literature
| S-EPMC4000852 | biostudies-literature
| S-EPMC7002611 | biostudies-literature
| S-EPMC4818741 | biostudies-other
| S-EPMC7224336 | biostudies-literature
| S-EPMC5036862 | biostudies-literature
| S-EPMC6767390 | biostudies-literature
| S-EPMC6292586 | biostudies-literature
| S-EPMC4998859 | biostudies-other
| S-EPMC4301848 | biostudies-literature