Quantitative Characterization of the T Cell Receptor Repertoire of Naive and Memory Subsets Using an Integrated Experimental and Computational Pipeline Which Is Robust, Economical, and Versatile.

ABSTRACT: The T cell receptor (TCR) repertoire can provide a personalized biomarker for infectious and non-infectious diseases. We describe a protocol for amplifying, sequencing, and analyzing TCRs which is robust, sensitive, and versatile. The key experimental step is ligation of a single-stranded oligonucleotide to the 3' end of the TCR cDNA. This allows amplification of all possible rearrangements using a single set of primers per locus. It also introduces a unique molecular identifier to label each starting cDNA molecule. This molecular identifier is used to correct for sequence errors and for effects of differential PCR amplification efficiency, thus producing more accurate measures of the true TCR frequency within the sample. This integrated experimental and computational pipeline is applied t

SUBMITTER: Oakes T 

PROVIDER: S-EPMC5643411 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature