Conserved nucleotide sequences at the 5' end of T cell receptor variable genes facilitate polymerase chain reaction amplification

Abstract

Alignment of all available nucleotide sequences of mouse and rat α/β T cell receptor (TcR) variable (V) regions revealed the presence of relatively conserved sequences at the 5' end of the V gene segments. Based on these conserved sequences, degenerate primers were developed for use in the polymerase chain reaction (PCR). The degenerate primers developed on the basis of the conserved sequences at the 5' end of rat and mouse V gene segments are expected to enable the amplification of all mouse and rat TcR α/β chain V regions. To test their applicability, the primers were used for the amplification of the V region of the TcR α/β expressed by rat T cell lines. After amplification, the TcR V regions expressed were cloned and sequenced. The Zla T cell line was shown to use the same TcR V gene segments (V_a2 and V_β8.2), as most other experimental allergic encephalomyelitis associated T cell lines, but had different D and J segments. In spite of these differences at the nucleotide level, a remarkable conservation of the amino acid sequence at the V_bTDβ_Jβ junction was found. Alignment of a large number of human V_α and V_β gene segments revealed the presence of similarly conserved sequences. Degenerate primers based on these conserved sequences enabled the amplification of TcR V regions of human T cell lines.