Evaluation of arrayed primer extension for TP53 mutation detection in breast and ovarian carcinomas

• Published in: BioTechniques Vol. 39; no. 5; pp. 755 - 761
• Main Authors: KRINGEN, Pedro, BERGAMASCHI, Anna, UNDLIEN DUE, Eldri, YUN WANG, TAGLIABUE, Elda, NESLAND, Jahn M, NEHMAN, Aune, TÖNISSON, Neeme, BÖRRESEN-DALE, Anne-Lise
• Format: Journal Article
• Language: English
• Published: Natick, MA Eaton 01.11.2005; Future Science Ltd
• Subjects: Alleles; Biological and medical sciences; Breast Neoplasms - genetics; Cell Line, Tumor; Codon; Diverse techniques; DNA Fragmentation; DNA Mutational Analysis - methods; DNA Primers - chemistry; DNA Primers - genetics; Electrophoresis; Female; Female genital diseases; Fundamental and applied biological sciences. Psychology; Genes, p53; Gynecology. Andrology. Obstetrics; Humans; Immunohistochemistry; Mammary gland diseases; Medical sciences; Molecular and cellular biology; Mutation; Oligonucleotide Array Sequence Analysis - methods; Oligonucleotides, Antisense - chemistry; Ovarian Neoplasms - genetics; Polymerase Chain Reaction; Sequence Analysis, DNA; Time Factors; Tumor Suppressor Protein p53 - genetics; Tumors; Array; Mammary gland diseases; Ovarian diseases; Breast carcinoma; Ovary carcinoma; TP53 Gene; Primer; Malignant tumor; Mutation; Detection; Female genital diseases
• ISSN: 0736-6205; 1940-9818
• DOI: 10.2144/000112000

Mutations in the tumor suppressor gene TP53 are associated with a wide range of different cancers and may have prognostic and therapeutic implications. Methods for rapid and sensitive detection of mutations in this gene are therefore required. In order to make screening more effective, a commercially available TP53 genotyping microarray from Asper Biotech has been constructed by arrayed primer extension (APEX). The present study is the first report that blindly evaluates the efficiency of the second generation APEX TP53 genotype chip outside the Asper laboratory and compares it to temporal temperature gradient electrophoresis (TTGE) and sequencing of TP53 for mutation detection in ovarian and breast cancer samples. All nucleotides in the TP53 gene from exon 2-9 are included on the chip by synthesis and application of sequence-specific oligonucleotides. The chip was validated by screening 48 breast and 11 ovarian cancer cases, all of which had previously been analyzed by TTGE and sequencing. APEX scored 17 of 20 sequence variants, missing one deletion, one insertion, and a missense mutation. Resequencing efficiency using APEX was 92% for both DNA strands and 99.5% for sense and/or antisense strand. We conclude that the APEX TP53 microarray is a robust, rapid, and comprehensive screening tool for sequence alterations in tumors.