title: Genotyping of the lactase-phlorizin hydrolase c/t-13910 polymorphism by means of a new rapid denaturing high-performance liquid chromatography-based assay in healthy subjects and colorectal cancer patients.
abstract: Adult-type hypolactasia results from the progressive decline of lactase-phlorizin hydrolase activity in enterocytes after weaning. Lactase nonpersistence may determine a primary lactose intolerance with reduced diary product consumption, which is possibly related to an increased risk of colon cancer. Recently, a genetic variant C/T(-13910) upstream of the lactase-phlorizin hydrolase (LCT) gene has been strongly correlated with the lactase persistence/nonpersistence trait in both family and case-control studies. The authors validate a denaturing high-performance liquid chromatography (dHPLC)-based assay versus conventional genotype sequencing in detecting the C/T(-13910) polymorphism of LCT and evaluate its prevalence in 2 different Italian geographical areas and in colorectal cancer patients. DNA samples of 157 healthy subjects and 124 colon cancer patients from Apulia and of 97 healthy subjects from Sardinia were evaluated for the C/T(-13910) polymorphism by dHPLC, sequencing, and restriction fragment length polymorphism (RFLP). Under optimized conditions, dHPLC was as sensitive as DNA sequencing and detected a new genetic variant (T/C(-13913)) in 2 individuals that was not identified by RFLP assay. Frequency of lactase nonpersistence genotype (C/C(-13910)) was similar in healthy subjects from 2 different Italian geographical areas and not increased in patients with colorectal cancer. The results indicate that the dHPLC method may be used as a rapid, noninvasive, and labor-saving screening tool for genotyping C/T(-13910) polymorphism, with high success, low cost, and reproducibility.
authors.affiliation: Research Laboratory and Department of Gastroenterology, Casa Sollievo della Sofferenza Hospital, IRCCS, San Giovanni Rotondo, Italy. a.piepoli@operapadrepio.it
authors.forename: Ada
authors.initials: A
authors.lastname: Piepoli
authors.forename: Enrico
authors.initials: E
authors.lastname: Schirru
authors.forename: Angela
authors.initials: A
authors.lastname: Mastrorilli
authors.forename: Annamaria
authors.initials: A
authors.lastname: Gentile
authors.forename: Rosa
authors.initials: R
authors.lastname: Cotugno
authors.forename: Michele
authors.initials: M
authors.lastname: Quitadamo
authors.forename: Antonio
authors.initials: A
authors.lastname: Merla
authors.forename: Mauro
authors.initials: M
authors.lastname: Congia
authors.forename: Paolo
authors.initials: P
authors.lastname: Usai Satta
authors.forename: Francesco
authors.initials: F
authors.lastname: Perri
mesh_terms: D000328:Adult; D001681:Biological Assay / Q000379:methods*; D016022:Case-Control Studies; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D015179:Colorectal Neoplasms / Q000201:enzymology / Q000235:genetics* / Q000473:pathology*; D005260:Female; D014644:Genetic Variation; D005838:Genotype; D006579:Heterozygote; D006720:Homozygote; D006801:Humans; D007558:Italy / Q000453:epidemiology; D043322:Lactase / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D010694:Lactase-Phlorizin Hydrolase / Q000235:genetics*; D007787:Lactose Intolerance / Q000201:enzymology / Q000235:genetics; D008297:Male; D008875:Middle Aged; D009367:Neoplasm Staging; D011110:Polymorphism, Genetic; D012150:Polymorphism, Restriction Fragment Length; D015995:Prevalence; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013997:Time Factors
chemical_list: D043322:Lactase; D010694:Lactase-Phlorizin Hydrolase