Polymerase chain reaction haplotyping using 3′ mismatches in the forward and reverse primers: application to the biallelic polymorphisms of tumor necrosis factor and lymphotoxin
Fanning, G.C., Bunce, M., Black, C.M. and Welsh, K.I. (1997) Polymerase chain reaction haplotyping using 3′ mismatches in the forward and reverse primers: application to the biallelic polymorphisms of tumor necrosis factor and lymphotoxin. Tissue Antigens, 50 (1). pp. 23-31.
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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 α (LT-α). 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-α 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-α 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-α. A total of 11 TNF-LT-α haplotypes were determined from apparent homozygous controls and statistical analysis.
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