Primer sets for cloning the human repertoire of T cell Receptor Variable regions

Department of Medical Sciences and Interdisciplinary Research Centre for Autoimmune Diseases, via Solaroli 17, 28100, Novara, Italy.
BMC Immunology (Impact Factor: 2.48). 09/2008; 9(1):50. DOI: 10.1186/1471-2172-9-50
Source: PubMed


Amplification and cloning of naïve T cell Receptor (TR) repertoires or antigen-specific TR is crucial to shape immune response and to develop immuno-based therapies. TR variable (V) regions are encoded by several genes that recombine during T cell development. The cloning of expressed genes as large diverse libraries from natural sources relies upon the availability of primers able to amplify as many V genes as possible.
Here, we present a list of primers computationally designed on all functional TR V and J genes listed in the IMGT, the ImMunoGeneTics information system. The list consists of unambiguous or degenerate primers suitable to theoretically amplify and clone the entire TR repertoire. We show that it is possible to selectively amplify and clone expressed TR V genes in one single RT-PCR step and from as little as 1000 cells.
This new primer set will facilitate the creation of more diverse TR libraries than has been possible using currently available primer sets.

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    • "First strand cDNA synthesis was primed with the specific primer TRBC-R4 (ctcaggcagtatctggagtcattgag) in the Superscript First-strand synthesis System (Invitrogen). To amplify the Vβ CDR3 region, PCR and nested PCR were performed with 17 different TRBV specific primers [18] and the reverse primer TRBC-R3 (tcaaacacagcgacctcgggtg). PCR products were cloned with the TOPO TA cloning kit (Invitrogen). "
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