Genotyping of the Lactase-Phlorizin Hydrolase −13910 Polymorphism by LightCycler PCR and Implications for the Diagnosis of Lactose Intolerance

Abstract

Background: Hypolactasia and lactose intolerance are common conditions worldwide. Hypolactasia seems to be strongly correlated with genotype C/C of the genetic variant C→T−13910 upstream of the lactase phlorizin hydrolase (LPH) gene. We developed a rapid genotyping assay for LPH C→T−13910 and investigated the relationship of positive lactose breath hydrogen test (LBHT) results suggesting lactose intolerance with LPH C→T−13910 genotype. Methods: Using automated DNA purification on the MagNA Pure LC and real-time PCR on the LightCycler, we examined samples from 220 individuals to estimate genotype frequencies; we then determined LPH C→T−13910 genotype in samples from 54 Caucasian patients with a positive LBHT result and symptoms of lactose intolerance. Results: Genotyping of 220 individuals revealed frequencies of 21.4%, 41.8%, and 36.8% for genotypes C/C, C/T, and T/T. Of the patients with positive LBHT results, only 50% had the C/C genotype suggestive of primary adult hypolactasia in our study population. The other patients had various degrees of secondary hypolactasia or symptoms of lactose intolerance. Patients with C/C genotype had a mean (SD) peak H2 increase in the LBHT [108 (58) ppm] that was significantly higher than in patients with the C/T [65 (54) ppm] and T/T [44 (34) ppm] genotypes. Conclusions: The new real-time PCR assay provides a rapid, labor-saving means for the genotyping of LPH C→T−13910. Use of the assay may assist in differentiating patients with primary hypolactasia from those with secondary hypolactasia and lactose intolerance, who may need further clinical examinations to diagnose their underlying primary diseases.

Rapid Genotyping of the Human Renin (REN) Gene by the LightCycler® Instrument: Identification of Unexpected Nucleotide Substitutions within the Selected Hybridization Probe Area

“…Several diagnostic assays have been developed on the LightCycler instrument, enabling the identification of genotypes by characteristic melting peaks and T m shifts [20][21][22]. Some of these well-established diagnostic assays have reported atypical melting peaks due to the presence of new nucleotide substitutions in the hybridization probe area beside those the assays were originally designed for.…”

Correlation between lactose absorption and the C/T-13910 and G/A-22018 mutations of the lactase-phlorizin hydrolase (LCT) gene in adult-type hypolactasia

“…Polymorphism of C/T-13910 has been used as a diagnostic predictor of adult-type hypolactasia in some European countries (5,23,24). However, this polymorphism has not been associated with lactase persistence in some African SubSaharan populations since several milkdrinking groups did not show the T-13910 allele.…”

Single nucleotide polymorphism C/T-13910, located upstream of the lactase gene, associated with adult-type hypolactasia: Validation for clinical practice

“…Samples showing a single band of 201 bp were classified as the CC genotype or a single band of 177 bp as the TT genotype; two bands of 201 bp and 177 bp represented the CT genotype. The CC genotype was previously associated with hypolactasia, and the CT and TT genotypes with normolactasia [3][4][5][6][7][8][9][10].…”