Table of Contents
ISRN Molecular Biology
Volume 2013 (2013), Article ID 451298, 4 pages
Research Article

Efficient IDUA Gene Mutation Detection with Combined Use of dHPLC and Dried Blood Samples

1Departamento de Genética Humana, Unidade I&D-P DLS, CGMJM, Instituto Nacional de Saúde Dr. Ricardo Jorge (INSA, IP), Pr. Pedro Nunes 88, 4099-028 Porto, Portugal
2Departamento de Genética Humana, Unidade de Tecnologia e Inovação (UTI), Instituto Nacional de Saúde Dr. Ricardo Jorge (INSA, IP), Avenida Padre Cruz, 1649-016 Lisboa, Portugal

Received 14 March 2013; Accepted 3 April 2013

Academic Editors: D. Bozon, E. Caffarelli, A. D. Hollenbach, B. L. Nielsen, and C.-H. Yuh

Copyright © 2013 Diogo Ribeiro et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


Objectives. Development of a simple mutation directed method in order to allow lowering the cost of mutation testing using an easily obtainable biological material. Assessment of the feasibility of such method was tested using a GC-rich amplicon. Design and Methods. A method of denaturing high-performance liquid chromatography (dHPLC) was improved and implemented as a technique for the detection of variants in exon 9 of the IDUA gene. The optimized method was tested in 500 genomic DNA samples obtained from dried blood spots (DBS). Results. With this dHPLC approach it was possible to detect different variants, including the common p.Trp402Ter mutation in the IDUA gene. The high GC content did not interfere with the resolution and reliability of this technique, and discrimination of G-C transversions was also achieved. Conclusion. This PCR-based dHPLC method is proved to be a rapid, a sensitive, and an excellent option for screening numerous samples obtained from DBS. Furthermore, it resulted in the consistent detection of clearly distinguishable profiles of the common p.Trp402Ter IDUA mutation with an advantageous balance of cost and technical requirements.