Polymerase Chain Reaction with Single-Sided Specificity: Analysis of T Cell Receptor δ Chain

Abstract

In the polymerase chain reaction (PCR), two specific oligonucleotide primers are used to amplify the sequences between them. However, this technique is not suitable for amplifying genes that encode molecules where the 5' portion of the sequences of interest is not known, such as the T cell receptor (TCR) or immunoglobulins. Because of this limitation, a novel technique, anchored polymerase chain reaction (A-PCR), was devised that requires sequence specificity only on the 3' end of the target fragment. It was used to analyze TCR delta chain mRNA's from human peripheral blood gamma delta T cells. Most of these cells had a V delta gene segment not previously described (V delta 3), and the delta chain junctional sequences formed a discrete subpopulation compared with those previously reported.

Immunophenotypic and immunogenotypic characteristics of TCRγδ+ T cell acute lymphoblastic leukemia

“…Deletion of nucleotides is probably caused by random exonucleolytic nibbling at the ends of most involved gene segments. 9,[20][21][22][23][24][25][26] Junctional regions are made up of D gene segments, N-region nucleotides, which are randomly inserted by the enzyme terminal deoxynucleotidyl transferase (TdT), 9,10,16,20,21,[23][24][25][26] and P-region nucleotides, ie maximally two palindromic nucleotides in juxtaposition to an untrimmed gene segment. 22 The junctional diversity of the TCRG and TCRD genes has been suggested to compensate for the limited combinatorial diversity of ␥␦ receptors.…”

Characterization and genomic sequence of the murine 60 kD Ro gene

“…An anchored polymerase chain reaction (PCR) was performed as previously described. 29 Briefly, liver and kidney poly (A) + RNA was isolated from Balb/c mice using standard procedures. Single strand cDNA was synthesized using Maloney murine leukemia virus reverse transcriptase (Promega, Madison, WI, USA).…”

Cloning and characterization of a complementary DNA for a thyroid hormone-responsive protein in mature rat cerebral tissue

“…Nucleotide sequence analysis showed that the clone contains a single long open reading frame, but lacked the initiation codon. The remaining coding sequence, along with part of the 5h untranslated region, was obtained by 5h rapid amplification of cDNA ends [20][21][22] (Gibco BRL, Gaithersburg, MD, U.S.A.).…”