Denaturing high‐performance liquid chromatography: A review

By W. Xiao, Peter J. Oefner. Human Mutation, 2001

700 citations recorded by the source corpus.

Denaturing high‐performance liquid chromatography (DHPLC) compares two or more chromosomes as a mixture of denatured and reannealed PCR amplicons, revealing the presence of a mutation by the differential retention of homo‐ and heteroduplex DNA on reversed‐phase chromatography supports under partial denaturation. Temperature determines sensitivity, and its optimum can be predicted by computation. Single‐nucleotide substitutions, deletions, and insertions have been detected successfully by on‐line UV or fluorescence monitoring within 2–3 minutes in unpurified amplicons as large as 1.5 Kb. Sensitivity and specificity of DHPLC consistently exceed 96%. These features and its low cost make DHPLC one of the most powerful tools for the re‐sequencing of the human and other genomes. Aside from its application to the mutational analysis of candidate genes, DHPLC has proven instrumental in elucidating human evolution and in the mapping of genes. Employing completely denaturing conditions, the utility of DHPLC has been extended to the genotyping of known polymorphisms by utilizing the ability of poly(styrene‐divinylbenzene) to resolve single‐stranded DNA molecules of identical size that differ in a single base. Under completely denaturing conditions, it is thus possible to resolve all possible base substitutions with the single exception of C→G transversions. Improvements in throughput became feasible with the recent introduction of monolithic poly(styrene‐divinylbenzene) capillaries that lend themselves to the fabrication of arrays connected to a multi‐color laser induced fluorescence scanner or a mass spectrometer. Hum Mutat 17:439–474, 2001. © 2001 Wiley‐Liss, Inc.

Subfields: Genetics, Molecular Biology

View Paper: https://www.semanticscholar.org/paper/83c3b34673da2be3796f5a4f8775254ca1ba37ba

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Citations in the corpus

Mutation of GABRA1 in an autosomal dominant form of juvenile myoclonic epilepsy (2002, methodology): “We amplified portions of GABRA1 by PCR and analyzed them by SSCP and dHPL”
Genetics of HUS: the impact of MCP, CFH, and IF mutations on clinical presentation, response to treatment, and outcome. (2006, unknown): “Treated episodes 57 (61) 23 (35) 6 (8) .”
The gene mutated in autosomal recessive polycystic kidney disease encodes a large, receptor-like protein (2002, methodology): “For the second strategy we used denaturing high-performance liquid chromatography (DHPLC), a rapid and sensitive method for detecting base-pair change”
Mutations in a novel gene lead to kidney tumors, lung wall defects, and benign tumors of the hair follicle in patients with the Birt-Hogg-Dubé syndrome. (2002, background): “The unique DHPLC 2-peak chroMutation analysis of the putative BHD gene reveals frameshift mutations which are predicted matographic profile was produced by separation of heteroduplexes, formed between wild-type and mutant allele amplicons, to truncate the protein The exon/intron structure of the BHD”
PTPN11 mutations in Noonan syndrome: molecular spectrum, genotype-phenotype correlation, and phenotypic heterogeneity. (2002, methodology): “Since this method is purported to have a sensitivity of 96%–100% under ideal conditions (Xiao and Oefner 2001), the true prevalence of point mutations in this cohort could be slightly higher.”
Consensus meeting for the management of gastrointestinal stromal tumors. Report of the GIST Consensus Conference of 20-21 March 2004, under the auspices of ESMO. (2005, methodology): “Denaturing high-performance liquid chromatography has recently been used to detect KIT and PDGFRa mutations in GIST [24–26], and is one of the most sensitive techniques for the detection of mutations [27].”