The application of real-time PCR to the analysis of T cell repertoires

Peter Wettstein, Michael Strausbauch, Terry Therneau, Nancy Borson

Research output: Contribution to journalArticle

13 Scopus citations

Abstract

The diversity of T-cell populations is determined by the spectrum of antigen-specific T-cell receptors (TCRs) that are heterodimers of α and β subunits encoded by rearranged combinations of variable (AV and BV), joining (AJ and BJ), and constant region genes (AC and BC). We have developed a novel approach for analysis of β transcript diversity in mice with a real-time PCR-based method that uses a matrix of BV- and BJ-specific primers to amplify 240 distinct BV-BJ combinations. Defined endpoints (Ct values) and dissociation curves are generated for each BV-BJ combination and the Ct values are consolidated in a matrix that characterizes the β transcript diversity of each RNA sample. Relative diversities of BV-BJ combinations in individual RNA samples are further described by estimates of scaled entropy. A skin allograft system was used to demonstrate that dissection of repertoires into 240 BV-BJ combinations increases efficiency of identifying and sequencing β transcripts that are overrepresented at inflammatory sites. These BV-BJ matrices should generate greater investigation in laboratory and clinical settings due to increased throughput, resolution and identification of overrepresented TCR transcripts.

Original languageEnglish (US)
Article numbere140
JournalNucleic acids research
Volume36
Issue number21
DOIs
StatePublished - 2008

ASJC Scopus subject areas

  • Genetics

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