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Detection of APC mutations by a yeast‐based protein truncation test (YPTT)
Author(s) -
Suzuki Takao,
Ishioka Chikashi,
Kato Satoshi,
Mitachi Yasushi,
Shimodaira Hideki,
Sakayori Masato,
Shimada Akira,
Asamura Mitsuo,
Kanamaru Ryunosuke
Publication year - 1998
Publication title -
genes, chromosomes and cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.754
H-Index - 119
eISSN - 1098-2264
pISSN - 1045-2257
DOI - 10.1002/(sici)1098-2264(199804)21:4<290::aid-gcc2>3.0.co;2-u
Subject(s) - frameshift mutation , nonsense mutation , biology , microbiology and biotechnology , adenomatous polyposis coli , gene , mutation , familial adenomatous polyposis , genetics , missense mutation , cancer , colorectal cancer
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. When 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. Genes Chromosomes Cancer 21:290–297, 1998. © 1998 Wiley‐Liss, Inc.