Denaturing High-Performance Liquid Chromatography and Sequence Analyses for MECP2 Mutations in Rett Syndrome

The detection of abnormal variations in a DNA sequence is a challenge for genetic research and clinical diagnostic applications. Among the different mutation detection methods currently available, DNA sequence analysis is largely considered the gold standard because it provides complete information about the nature and location of a particular sequence variant. The development of polymerase chain reaction (PCR)based direct sequencing protocols using automated fluorescence detection systems has greatly facilitated DNA sequencing. In spite of numerous improvements in sequencing technologies, however, full-scale DNA sequence analysis remains relatively costly and labor-intensive in many laboratory settings. Unknown sequence variants in a region of interest may also be detected using a variety of mutation-scanning methods (1 ). These scanning methods have inherent strengths and limitations, and typically require confirmatory sequence analysis to identify the DNA alteration as a mutation or a non-disease-associated variant. Technical strengths may include speed, ease of use, and lower assay costs, but the critical limitation for most scanning methods involves a suboptimal mutation-detection rate.