Detection of APC mutations by a yeast-based protein truncation test (YPTT)

Takao Suzuki, Chikashi Ishioka, Satoshi Kato, Yasushi Mitachi, Hideki Shimodaira, Masato Sakayori, Akira Shimada, Mitsuo Asamura, Ryunosuke Kanamaru

Research output: Contribution to journalArticlepeer-review

9 Citations (Scopus)

Abstract

APC gene mutations play a role in the initiation step of colorectal carcinogenesis in both familial adenomatous polyposis (FAP) and non-FAP patients. Almost all of the APC mutations are nonsense or frameshift mutations, which truncate the APC protein and are thought to inactivate normal APC function. We show a novel method for detecting nonsense and frameshift APC gene mutations by using Saccharomyces cerevisiae. Polymerase chain reaction (PCR)-amplified APC fragments are cloned directly into yeast expression vectors in vivo, and the yeast expresses a hemagglutinin epitope (HA)-tagged APC peptide. Widen an APC fragment contains a nonsense or frameshift mutation, HA-tagged truncating APC peptide can be detected by Western blotting using an anti-HA antibody. We identified both germ-line and somatic APC mutations in patients with FAP and non-FAP colorectal tumors, respectively. This method, called the yeast-based protein truncation test (YPTT), is simple and fairly cheap, and it can be applied to any genes that are inactivated by protein truncating mutations.

Original languageEnglish
Pages (from-to)290-297
Number of pages8
JournalGenes Chromosomes and Cancer
Volume21
Issue number4
DOIs
Publication statusPublished - 1998
Externally publishedYes

ASJC Scopus subject areas

  • Genetics
  • Cancer Research

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