Novel Hairpin-Shaped Primer Assay To Study the Association of the −44 Single-Nucleotide Polymorphism of theDEFB1Gene with Early-Onset Periodontal Disease

Abstract

A powerful, cost-effective new method for studying single-nucleotide polymorphisms (SNPs) is described. This method is based on the use of hairpin-shaped primers (HP), which give a sensitive and specific PCR amplification of each specific allele, without the use of costly fluorophore-labeled probes and any post-PCR manipulation. The amplification is monitored in real-time using SYBR Green I dye and takes only 2 h to yield results. The HP assay has a simple design and utilizes a conventional real-time PCR apparatus. The ؊44 C3G transversion in the DEFB1 gene (which encodes human ␤-defensin 1) has been previously associated with Candida carriage in oral epithelia. In this study, we analyzed the association between early-onset periodontal disease (EOP) and the ؊44 SNP. We used an HP assay to study the distribution of the ؊44 SNP in 264 human DNAs obtained from two cohorts of EOP patients and healthy controls from different ethnic backgrounds. The results indicate that the ؊44 SNP has a similar distribution between EOP and healthy patients, suggesting that it is not associated with the disease.

A 3′ UTR transition within DEFB1 is associated with chronic and aggressive periodontitis

“…The robustness of these data was further supported by a separate analysis of the two parodontopathic distinct forms: CP and AgP. Both independent case populations differed only marginally in terms of allele frequencies and contributed in a Earlier studies on periodontitis investigated associations of SNP G-20A (rs11362) 20 and C-44G (rs1800972) 19 with patients with severe CP and those with early onset periodontitis (currently most likely classified as AgP), respectively. These studies were of much smaller sample size than this study and observed no disease association.…”

“…In earlier studies, the promoter SNPs: G-20A (rs11362) 20 and C-44G (rs1800972) 19 were subjected to candidate gene association studies in patients with severe CP and early onset periodontitis. In these studies, no association with periodontitis was observed.…”

“…In addition, we analyzed associations of the 5 0 UTR SNP G-20A (rs11362), C-44G (rs1800972) and G-52A (rs1799946), two of which have been earlier subjected to candidate gene association studies in patients with severe CP and early onset periodontitis (see Table 2). 19,20 Before adjustment for covariates: smoking, diabetes and the potential confounder gender using logistic regression analysis, five haplotype-tagging SNPs (rs1047031, rs2293958, rs2980930, rs5743401 and rs5743402) and the promoter SNP rs1800972 gave evidence for association with periodontitis (Supplementary Table S1). We next adjusted for covariates: smoking, diabetes and gender using logistic regression analysis.…”

Human β-Defensin-1, a Potential Chromosome 8p Tumor Suppressor: Control of Transcription and Induction of Apoptosis in Renal Cell Carcinoma

“…The ''mutant'' promoter contains the second most common base (C) at position À688 and (G) at À44. These variants are prevalent in the population (1,5). Promoter deletion constructs were made to include a variable amount of upstream sequence.…”

“…Defensins are classified into a-defensins, h-defensins, and u-defensins according to the size and binding patterns of disulfide bonds within mature peptides. To date, six h-defensins have been identified and mapped to a locus on chromosome 8p, an area that contains at least three separate tumor suppressor genes inactivated during the progression of human malignancies, including prostate and renal cancers (1)(2)(3). Previous studies have revealed that deletion of chromosome 8p is the most common genetic alteration in prostate cancer and the second most common genomic event in bladder and renal cancers (4).…”