Polymerase chain reaction haplotyping using 3′ mismatches in the forward and reverse primers: application to the biallelic polymorphisms of tumor necrosis factor and lymphotoxin α
- 1 July 1997
- journal article
- Published by Wiley in Tissue Antigens
- Vol. 50 (1) , 23-31
- https://doi.org/10.1111/j.1399-0039.1997.tb02829.x
Abstract
A polymerase chain reaction with sequence-specific primers (PCR-SSP) system that operates under identical conditions to HLA phototyping was devised for characterizing polymorphisms in tumor necrosis factor (TNF) and lymphotoxin alpha (LT-alpha). Mismatches at the 3' end were incorporated into the forward and reverse primers of each PCR so as to unequivocally establish the cis/trans status between the biallelic sites. Three previously described biallelic polymorphisms in TNF and three in LT-alpha were characterized in a 24-reaction PCR-SSP system. The method was used to genotype 20 cell lines and 201 HLA class I and II typed controls from the United Kingdom at the TNF and LT-alpha loci. Population frequencies of TNF haplotypes were determined as was linkage disequilibrium with HLA-A, B, Cw, DRB1 and DQB1 loci. In each gene there were 8 theoretical polymorphic combinations; 4 were observed in TNF and 4 in LT-alpha. A total of 11 TNF-LT-alpha haplotypes were determined from apparent homozygous controls and statistical analysis.Keywords
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