Genotyping of TNF Gene Polymorphisms

Two TNF gene polymorphisms, previously associated with tuberculosis in European, Mexican American, Hispanic, and Latino populations, were selected [19 (link), 28 (link)–31 (link)]. TNF –308G/A and –238G/A gene polymorphisms were genotyped by polymerase chain reaction and restriction fragment length polymorphisms (PCR-RFLP) [32 (link)]. The –308G/A homozygous AA genotype is identified by a 107 bp fragment, the GG genotype has a combination of 87 and 20 bp bands, and the heterozygous GA has three bands (107, 87, 20 bp). The –238 AA polymorphic genotype is identified by a 152 bp fragment, the GG wild type has two bands (133, 19 bp), and the heterozygous GA has three fragments. Genotypes were confirmed by Sanger sequencing with an ABI Prism 3130 genetic analyser and ABI Prism BigDye Terminator^® v3.1 (Applied Biosystems, USA); each genotype was confirmed twice in a bidirectional way by PCR (Figure 1).