Identification of novel human T-cell receptor V beta gene segments by the anchored-polymerase chain reaction.
We have studied the diversity of the expressed human T-cell receptor (TCR) beta-chain repertoire by analysis of mRNA from unstimulated peripheral blood T-cells. The anchored-polymerase chain reaction (PCR) was used to isolate TCRB transcripts. Of 20 full or near full-length functional transcripts sequenced, two were novel TCRVB gene segments. They have strong sequence similarities to the known TCRBV5, and 8 subfamilies. Southern blot analysis and sequence-specific oligonucleotide hybridization confirmed: a) that these sequences are present in genomic DNA; b) their relationship to the known TCRBV families. A TCRBV sequence similar to a recently identified novel TCRBV24 subfamily was also found. We show by southern blotting that this sequence forms a single member subfamily, and by deletion analysis of T-cell lines, we have mapped this sequence to lie between the genes which encode the TCRBV8.1 and TCRBV5.3 gene segments. The results show that the anchored PCR is a powerful tool in the analysis of the TCR repertoire, which may contain more V gene segments than previously defined.
Lunardi
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Department of Medicine, Royal Postgraduate Medical School, London, UK.
Marguerie
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D000595:Amino Acid Sequence; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000235:genetics*
D016693:Receptors, Antigen, T-Cell, alpha-beta; D004247:DNA