This Article Full Text Full Text (PDF) Purchase Article View Shopping Cart Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Vaughn, C. P. Articles by Elenitoba-Johnson, K. S. J. Search for Related Content PubMed PubMed Citation Articles by Vaughn, C. P. Articles by Elenitoba-Johnson, K. S. J.

JMD 2004, Vol. 6, No. 3
Copyright © 2004 American Society for Investigative Pathology & Association for Molecular Pathology

High-Resolution Melting Analysis for Detection of Internal Tandem Duplications

Cecily P. Vaughn^* and Kojo S. J. Elenitoba-Johnson^*

From the Associated Regional and University Pathologists Institute for Clinical and Experimental Pathology * and the Department of Pathology, University of Utah School of Medicine, Salt Lake City, Utah

High-resolution melting analysis (HRMA) is a recently introduced closed-tube fluorescence-based method for rapid mutation screening and detection. However, all of the targets by which this technique has been validated thus far have had single-base substitutions, deletions, or similarly small mutational deviations from the wild-type sequence. In the current study, we sought to determine the feasibility of utilization of HRMA for the detection of larger sequence aberrations, using internal tandem duplications (ITD) in the juxtamembrane domain of the FLT3 gene as a model system. This gene is important in the growth and differentiation of hematopoietic progenitors and ITDs in this gene have been identified in a subset of poor-prognosis acute myelogenous leukemias (AML). DNA extracted from 62 AML samples was analyzed on a prototype high-resolution melting instrument. The samples interrogated for the FLT3 ITDs were subjected to post-amplification denaturation with frequent and regular fluorescence acquisition. The fluorescence versus temperature melting graphs generated were analyzed for deviation from the profiles reproducibly obtained for the wild-type samples. Results by HRMA were compared to results obtained using capillary electrophoresis-based fragment analysis, temperature gradient capillary electrophoresis detection, and sequencing of ITDs. FLT3 ITDs were detected in 13 of 62 AML samples with 100% concordance between the detection methods. This study demonstrates the utility of HRMA to rapidly and accurately screen samples for the presence of large sequence aberrations including FLT3 ITDs.

This article has been cited by other articles:

				A. J. Stephens, J. Inman-Bamber, P. M. Giffard, and F. Huygens High-Resolution Melting Analysis of the spa Repeat Region of Staphylococcus aureus Clin. Chem., February 1, 2008; 54(2): 432 - 436. [Abstract] [Full Text] [PDF]

				M. L. Kennerson, T. Warburton, E. Nelis, M. Brewer, P. Polly, P. De Jonghe, V. Timmerman, and G. A. Nicholson Mutation Scanning the GJB1 Gene with High-Resolution Melting Analysis: Implications for Mutation Scanning of Genes for Charcot-Marie-Tooth Disease Clin. Chem., February 1, 2007; 53(2): 349 - 352. [Abstract] [Full Text] [PDF]

				J.-C. Cheng, C.-L. Huang, C.-C. Lin, C.-C. Chen, Y.-C. Chang, S.-S. Chang, and C.-P. Tseng Rapid Detection and Identification of Clinically Important Bacteria by High-Resolution Melting Analysis after Broad-Range Ribosomal RNA Real-Time PCR Clin. Chem., November 1, 2006; 52(11): 1997 - 2004. [Abstract] [Full Text] [PDF]