Long-range PCR facilitates the identification of PMS2-specific mutations

• Published in: Human mutation Vol. 27; no. 5; pp. 490 - 495
• Main Authors: Clendenning, Mark, Hampel, Heather, LaJeunesse, Jennifer, Lindblom, Annika, Lockman, Jan, Nilbert, Mef, Senter, Leigha, Sotamaa, Kaisa, de la Chapelle, Albert
• Format: Journal Article
• Language: English
• Published: Hoboken Wiley Subscription Services, Inc., A Wiley Company 01-05-2006; Hindawi Limited
• Subjects: Adaptor Proteins, Signal Transducing; Adenosine Triphosphatases - genetics; Basic Medicine; Carrier Proteins - genetics; Colorectal Neoplasms, Hereditary Nonpolyposis - diagnosis; Colorectal Neoplasms, Hereditary Nonpolyposis - genetics; DNA Mutational Analysis - methods; DNA Repair Enzymes - genetics; DNA-Binding Proteins - genetics; Genetic Predisposition to Disease; Genetic Testing; Humans; Lynch syndrome; Medical and Health Sciences; Medical Genetics; Medicin och hälsovetenskap; Medicinsk genetik; Medicinska och farmaceutiska grundvetenskaper; Mismatch Repair Endonuclease PMS2; Mutation; MutL Protein Homolog 1; Nuclear Proteins - genetics; PMS2; Polymerase Chain Reaction - methods; polymorphisms; Pseudogenes
• ISSN: 1059-7794; 1098-1004; 1098-1004
• DOI: 10.1002/humu.20318

Mutations within the DNA mismatch repair gene, “postmeiotic segregation increased 2” (PMS2), have been associated with a predisposition to hereditary nonpolyposis colorectal cancer (HNPCC; Lynch syndrome). The presence of a large family of highly homologous PMS2 pseudogenes has made previous attempts to sequence PMS2 very difficult. Here, we describe a novel method that utilizes long‐range PCR as a way to preferentially amplify PMS2 and not the pseudogenes. A second, exon‐specific, amplification from diluted long‐range products enables us to obtain a clean sequence that shows no evidence of pseudogene contamination. This method has been used to screen a cohort of patients whose tumors were negative for the PMS2 protein by immunohistochemistry and had not shown any mutations within the MLH1 gene. Sequencing of the PMS2 gene from 30 colorectal and 11 endometrial cancer patients identified 10 novel sequence changes as well as 17 sequence changes that had previously been identified. In total, putative pathologic mutations were detected in 11 of the 41 families. Among these were five novel mutations, c.705+1G>T, c.736_741del6ins11, c.862_863del, c.1688G>T, and c.2007–1G>A. We conclude that PMS2 mutation detection in selected Lynch syndrome and Lynch syndrome‐like patients is both feasible and desirable. Hum Mutat 27(5), 490–495, 2006. Published 2006 Wiley‐Liss, Inc.