Sequence-Specific Primer Polymerase Chain Reaction Genotyping of the Human IL33 Polymorphism rs1929992 (T > C) with...

Single-nucleotide polymorphisms (SNPs) may influence gene expression and contribute to interindividual variability in immune responses and disease susceptibility. Interleukin-33 (IL33), a cytokine involved in inflammation, immune regulation, epithelial barrier homeostasis, and tissue remodeling, has been implicated in allergic, autoimmune, inflammatory, and neoplastic disorders. The IL33 rs1929992 (T > C) polymorphism, located in intron 3, has been associated with altered IL33 expression, with the T allele linked to increased cytokine production compared with the C allele. This article describes a standardized sequence-specific primer polymerase chain reaction (PCR-SSP) protocol for genotyping the human IL33 rs1929992 polymorphism, with validation by Sanger sequencing. The protocol includes optimization of annealing temperature, MgCl₂ concentration, primer balance, deoxynucleotide triphosphates (dNTPs), and DNA input to ensure amplification specificity and reproducibility. Allele-specific amplification is achieved using forward primers specific for the T and C alleles in independent reactions and analyzed by agarose gel electrophoresis, with the human growth hormone (HGH) gene used as an internal control. This protocol provides a simple, reproducible, and accessible method for rs1929992 genotyping, suitable for population-based and genetic association studies involving immune-mediated and inflammatory diseases. © 2026 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: PCR-SSP genotyping of IL33 rs1929992 Support Protocol 1: Extraction of genomic DNA by salting-out.