NF2 mutation screening by denaturing high-performance liquid chromatography and high-resolution melting analysis. 2008

Roberta Sestini, and Aldesia Provenzano, and Costanza Bacci, and Claudio Orlando, and Maurizio Genuardi, and Laura Papi
Department of Clinical Physiopathology, Medical Genetics Unit, University of Florence, Firenze, Italy. r.sestini@dfc.unifi.it

Neurofibromatosis type 2 (NF2) is an autosomal-dominant disorder caused by mutations in the NF2 gene and predisposing to the development of nervous system. Identification of germline mutations is essential to provide appropriate genetic counseling in NF2 patients, but it represents an extremely challenging task because the vast majority of mutations are unique and spread over the entire coding sequence. Moreover, about 30% of de novo patients are indeed mosaic, and direct sequencing can undetect mutated alleles present in a minority of cells. As most screening techniques do not meet the requirements for efficient NF2 testing, we have developed a semi-automated denaturing high-performance liquid chromatography (DHPLC) method for point mutation detection combined with a multiplex ligation-dependent probe amplification approach to screen for gene rearrangements. In addition, we have evaluated high-resolution melting analysis (HRMA) as an exon scanning procedure to identify point mutations in the NF2 gene. The results obtained in 92 NF2 patients expand the NF2 mutational spectrum and indicate DHPLC and HRMA as good systems to screen for point mutations in diseases with a heterogeneous spectrum of alterations.

UI MeSH Term Description Entries
D009464 Neuroma, Acoustic A benign SCHWANNOMA of the eighth cranial nerve (VESTIBULOCOCHLEAR NERVE), mostly arising from the vestibular branch (VESTIBULAR NERVE) during the fifth or sixth decade of life. Clinical manifestations include HEARING LOSS; HEADACHE; VERTIGO; TINNITUS; and FACIAL PAIN. Bilateral acoustic neuromas are associated with NEUROFIBROMATOSIS 2. (From Adams et al., Principles of Neurology, 6th ed, p673) Acoustic Neuroma,Melanocytic Vestibular Schwannoma,Schwannoma, Acoustic,Schwannoma, Vestibular,Acoustic Neuroma, Cerebellopontine Angle,Acoustic Tumor,Angle Tumor,Cerebellopontine Angle Acoustic Neuroma,Cerebellopontine Angle Tumor,Neurilemmoma, Acoustic,Neurilemoma, Acoustic,Neurinoma of the Acoustic Nerve,Neurinoma, Acoustic,Neuroma, Acoustic, Unilateral,Vestibular Schwannoma,Acoustic Neurilemmoma,Acoustic Neurilemmomas,Acoustic Neurilemoma,Acoustic Neurilemomas,Acoustic Neurinoma,Acoustic Neurinomas,Acoustic Neuromas,Acoustic Schwannoma,Acoustic Schwannomas,Acoustic Tumors,Angle Tumor, Cerebellopontine,Angle Tumors,Angle Tumors, Cerebellopontine,Cerebellopontine Angle Tumors,Melanocytic Vestibular Schwannomas,Neurilemmomas, Acoustic,Neurilemomas, Acoustic,Neurinomas, Acoustic,Neuromas, Acoustic,Schwannoma, Melanocytic Vestibular,Schwannomas, Acoustic,Schwannomas, Melanocytic Vestibular,Schwannomas, Vestibular,Tumor, Acoustic,Tumor, Angle,Tumor, Cerebellopontine Angle,Tumors, Acoustic,Tumors, Angle,Tumors, Cerebellopontine Angle,Vestibular Schwannoma, Melanocytic,Vestibular Schwannomas,Vestibular Schwannomas, Melanocytic
D009691 Nucleic Acid Denaturation Disruption of the secondary structure of nucleic acids by heat, extreme pH or chemical treatment. Double strand DNA is "melted" by dissociation of the non-covalent hydrogen bonds and hydrophobic interactions. Denatured DNA appears to be a single-stranded flexible structure. The effects of denaturation on RNA are similar though less pronounced and largely reversible. DNA Denaturation,DNA Melting,RNA Denaturation,Acid Denaturation, Nucleic,Denaturation, DNA,Denaturation, Nucleic Acid,Denaturation, RNA,Nucleic Acid Denaturations
D002851 Chromatography, High Pressure Liquid Liquid chromatographic techniques which feature high inlet pressures, high sensitivity, and high speed. Chromatography, High Performance Liquid,Chromatography, High Speed Liquid,Chromatography, Liquid, High Pressure,HPLC,High Performance Liquid Chromatography,High-Performance Liquid Chromatography,UPLC,Ultra Performance Liquid Chromatography,Chromatography, High-Performance Liquid,High-Performance Liquid Chromatographies,Liquid Chromatography, High-Performance
D004252 DNA Mutational Analysis Biochemical identification of mutational changes in a nucleotide sequence. Mutational Analysis, DNA,Analysis, DNA Mutational,Analyses, DNA Mutational,DNA Mutational Analyses,Mutational Analyses, DNA
D005091 Exons The parts of a transcript of a split GENE remaining after the INTRONS are removed. They are spliced together to become a MESSENGER RNA or other functional RNA. Mini-Exon,Exon,Mini Exon,Mini-Exons
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D012680 Sensitivity and Specificity Binary classification measures to assess test results. Sensitivity or recall rate is the proportion of true positives. Specificity is the probability of correctly determining the absence of a condition. (From Last, Dictionary of Epidemiology, 2d ed) Specificity,Sensitivity,Specificity and Sensitivity
D016133 Polymerase Chain Reaction In vitro method for producing large amounts of specific DNA or RNA fragments of defined length and sequence from small amounts of short oligonucleotide flanking sequences (primers). The essential steps include thermal denaturation of the double-stranded target molecules, annealing of the primers to their complementary sequences, and extension of the annealed primers by enzymatic synthesis with DNA polymerase. The reaction is efficient, specific, and extremely sensitive. Uses for the reaction include disease diagnosis, detection of difficult-to-isolate pathogens, mutation analysis, genetic testing, DNA sequencing, and analyzing evolutionary relationships. Anchored PCR,Inverse PCR,Nested PCR,PCR,Anchored Polymerase Chain Reaction,Inverse Polymerase Chain Reaction,Nested Polymerase Chain Reaction,PCR, Anchored,PCR, Inverse,PCR, Nested,Polymerase Chain Reactions,Reaction, Polymerase Chain,Reactions, Polymerase Chain
D016515 Genes, Neurofibromatosis 2 Tumor suppressor genes located on the long arm of human chromosome 22. Mutation or loss of these genes causes NEUROFIBROMATOSIS 2. Genes, nf2,Neurofibromatosis 2 Genes,nf2 Genes,Genes, nf 2,Gene, Neurofibromatosis 2,Gene, nf 2,Gene, nf2,Neurofibromatosis 2 Gene,nf 2 Gene,nf 2 Genes,nf2 Gene
D016518 Neurofibromatosis 2 An autosomal dominant disorder characterized by a high incidence of bilateral acoustic neuromas as well as schwannomas (NEURILEMMOMA) of other cranial and peripheral nerves, and other benign intracranial tumors including meningiomas, ependymomas, spinal neurofibromas, and gliomas. The disease has been linked to mutations of the NF2 gene (GENES, NEUROFIBROMATOSIS 2) on chromosome 22 (22q12) and usually presents clinically in the first or second decade of life. Neurofibromatosis, Acoustic, Bilateral,Neurofibromatosis, Central, NF2,Neuroma, Acoustic, Bilateral,Schwannoma, Acoustic, Bilateral,Acoustic Neurinoma, Bilateral,Acoustic Schwannomas, Bilateral,Bilateral Acoustic Neurofibromatosis,Central Neurofibromatosis,Familial Acoustic Neuromas,NF2 (Neurofibromatosis 2),Neurofibromatosis II,Neurofibromatosis Type 2,Neurofibromatosis Type II,Neurofibromatosis, Central NF2,Neurofibromatosis, Central, NF 2,Neurofibromatosis, Type 2,Neurofibromatosis, Type II,Neurofibromatosis, central type,Acoustic Neurinomas, Bilateral,Acoustic Neurofibromatoses, Bilateral,Acoustic Neurofibromatosis, Bilateral,Acoustic Neuroma, Familial,Acoustic Neuromas, Familial,Acoustic Schwannoma, Bilateral,Bilateral Acoustic Neurinoma,Bilateral Acoustic Neurinomas,Bilateral Acoustic Neurofibromatoses,Bilateral Acoustic Schwannoma,Bilateral Acoustic Schwannomas,Central NF2 Neurofibromatoses,Central NF2 Neurofibromatosis,Central Neurofibromatoses,Familial Acoustic Neuroma,NF2s (Neurofibromatosis 2),Neurinoma, Bilateral Acoustic,Neurinomas, Bilateral Acoustic,Neurofibromatoses, Bilateral Acoustic,Neurofibromatoses, Central,Neurofibromatoses, Central NF2,Neurofibromatoses, Type 2,Neurofibromatoses, Type II,Neurofibromatosis IIs,Neurofibromatosis, Bilateral Acoustic,Neurofibromatosis, Central,Neuroma, Familial Acoustic,Neuromas, Familial Acoustic,Schwannoma, Bilateral Acoustic,Schwannomas, Bilateral Acoustic,Type 2 Neurofibromatoses,Type 2 Neurofibromatosis,Type II Neurofibromatoses,Type II Neurofibromatosis

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