Rapid detection of 3500Q and 3531 mutations and Mspl polymorphism in exon 26 at the apolipoprotein B gene

Several environmental and genetic factors are associated with high levels o f cholesterol. Hypercholesterolemia is the main phenotype of Familial Defec tive Apolipoprotein B and Familial Hypercholesterolemia that are caused by mutations at the apolipoprotein (apo) B and LDL receptor genes, respectivel y. Identification of the specific genetic alteration associated with hyperc holesterolemia is an important issue in clinical diagnosis of high risk for CAD. Apo B gene. mutations and polymorphisms are usually screened by SSCP, DGGE, and heteroduplex, which must be confirmed by DNA sequencing or by di rect detection using PCR techniques, In this study, we have optimized a PCR -RFLP procedure for identification of 3500Q and 3531 mutations and Mspl pol ymorphism at the apo B gene. The technique can be performed in a single rea ction, using the restriction endonuclease Mspl for simultaneous detection o f 3500Q mutation and Mspl polymorphism, and Nsil for detection of 3531 muta tion. The procedure was validated by analysis of control DNA samples from i ndividuals carrying these mutations. Screening of 186 Brazilian hypercholes terolemic individuals showed that the frequency of the M-allele (7.8%) of M spl polymorphism was similar to that found in other individuals with CAD. H owever, neither 3500Q nor 3531 mutations were detected in this group. In co nclusion, this procedure is simple and rapid, being easily introduced in cl inical laboratories for direct detection of the more frequent mutations at the apo B gene associated with hypercholesterolemia. J. Clin. Lab. Anal. 15 :35-39, 2001. (C) 2001 Wiley-Liss, Inc.