Enhanced detection of TP53 mutations using a GC-clamp in denaturing high performance liquid chromatography

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6 Citations (Scopus)

Abstract

Although denaturing gradient gel electrophoresis (DGGE) is a highly effective technique for screening for TP53 mutations, the use of denaturing high performance liquid chromatography (DHPLC) is a growing methodology. This report describes a comparison between DHPLC and DGGE in the detection of TP53 mutations in hematopoietic cell lines and lymphomas. In addition, the improved effectiveness of guanine cytosine (GC)-clamped DHPLC for TP53 screening is detailed. Thirty DNA samples with known TP53 mutations in the hotspot region of codons 5-8, previously identified by DGGE, were analyzed by DHPLC. We found 100% concordance in mutation detection by DHPLC with DGGE. Similar to the improved efficacy observed in DGGE, the addition of 40 nucleotide GC-clamps composed of guanine and cytosine bases at one end of the product enhanced the detection of a mutation pattern by DHPLC. DHPLC of GC-clamped products is a viable and faster alternative method for screening TP53 mutations.

Original languageEnglish (US)
Pages (from-to)32-37
Number of pages6
JournalDiagnostic Molecular Pathology
Volume16
Issue number1
DOIs
StatePublished - Mar 1 2007

Fingerprint

Cytosine
Guanine
Denaturing Gradient Gel Electrophoresis
High Pressure Liquid Chromatography
Mutation
Cytosine Nucleotides
Guanine Nucleotides
Codon
Lymphoma
Cell Line
DNA

Keywords

  • DGGE
  • Denaturing high performance liquid chromatography
  • GC-clamp
  • Mutation
  • TP53

ASJC Scopus subject areas

  • Pathology and Forensic Medicine
  • Molecular Biology
  • Cell Biology

Cite this

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title = "Enhanced detection of TP53 mutations using a GC-clamp in denaturing high performance liquid chromatography",
abstract = "Although denaturing gradient gel electrophoresis (DGGE) is a highly effective technique for screening for TP53 mutations, the use of denaturing high performance liquid chromatography (DHPLC) is a growing methodology. This report describes a comparison between DHPLC and DGGE in the detection of TP53 mutations in hematopoietic cell lines and lymphomas. In addition, the improved effectiveness of guanine cytosine (GC)-clamped DHPLC for TP53 screening is detailed. Thirty DNA samples with known TP53 mutations in the hotspot region of codons 5-8, previously identified by DGGE, were analyzed by DHPLC. We found 100{\%} concordance in mutation detection by DHPLC with DGGE. Similar to the improved efficacy observed in DGGE, the addition of 40 nucleotide GC-clamps composed of guanine and cytosine bases at one end of the product enhanced the detection of a mutation pattern by DHPLC. DHPLC of GC-clamped products is a viable and faster alternative method for screening TP53 mutations.",
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