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Titolo:
Optimal temperature selection for mutation detection by denaturing HPLC and comparison to single-stranded conformation polymorphism and heteroduplex analysis
Autore:
Jones, AC; Austin, J; Hansen, N; Hoogendoorn, B; Oefner, PJ; Cheadle, JP; ODonovan, MC;
Indirizzi:
Univ Wales Coll Med, Dept Psychol Med, Cardiff CF4 4XN, S Glam, Wales UnivWales Coll Med Cardiff S Glam Wales CF4 4XN f CF4 4XN, S Glam, Wales Univ Wales Coll Med, Dept Med Genet, Cardiff CF4 4XN, S Glam, Wales Univ Wales Coll Med Cardiff S Glam Wales CF4 4XN f CF4 4XN, S Glam, Wales
Titolo Testata:
CLINICAL CHEMISTRY
fascicolo: 8, volume: 45, anno: 1999,
parte:, 1
pagine: 1133 - 1140
SICI:
0009-9147(199908)45:8<1133:OTSFMD>2.0.ZU;2-R
Fonte:
ISI
Lingua:
ENG
Soggetto:
GRADIENT GEL-ELECTROPHORESIS; PERFORMANCE LIQUID-CHROMATOGRAPHY; THERMAL-DENATURATION; DNA; PROFILES;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
15
Recensione:
Indirizzi per estratti:
Indirizzo: O'Donovan, MC Univ Wales Coll Med, Dept Psychol Med, Heath Pk, Cardiff CF44XN, S Glam, Wales Univ Wales Coll Med Heath Pk Cardiff S Glam Wales CF4 4XN es
Citazione:
A.C. Jones et al., "Optimal temperature selection for mutation detection by denaturing HPLC and comparison to single-stranded conformation polymorphism and heteroduplex analysis", CLIN CHEM, 45(8), 1999, pp. 1133-1140

Abstract

Background: Denaturing HPLC (DHPLC) is a semiautomated method for detecting unknown DNA sequence variants. The sensitivity of the method is dependenton the temperature at which the analysis is undertaken, the selection of which is dependent on operator experience. To circumvent this, software has been developed for predicting the optimal temperature for DHPLC analysis. We examined the utility of this software. Methods: To maximize the relevance of our data for other investigators, wehave screened 42 different amplimers from CFTR, TSC1, and TSC2. The samples consisted of 103 unique sequence heterozygotes and 126 wild-type homozygous controls. Results: At the temperature recommended by the software, 96% (99 of 103) of heterozygotes and all of the wild-type controls were correctly classified. This compares favorably with sensitivities of 85% for single-stranded conformation polymorphism and 82% for gel-based heteroduplex analyses of the same fragments. Conclusions: Software-optimized DHPLC is a highly sensitive method for mutation detection. However, where sensitivity >96% is required, our data suggest that in addition to the recommended temperature, fragments should also be run at the recommended temperature plus 2 degrees C. (C) 1999 American Association for Clinical Chemistry.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 20/01/21 alle ore 15:29:04