Long Polymerase Chain Reaction-Based Fluorescence in Situ Hybridization Analysis of Female Carriers of X-Linked Chronic Granulomatous Disease Deletions

Kelly Claire Simon^*, Deborah Noack, Julie Rae, John Curnutte, Shireen Sarraf^*, Valentin Kolev and Jan K. Blancato^*

From the Lombardi Cancer Center, * Georgetown University Medical Center, Washington, District of Columbia; the Department of Obstetrics and Gynecology, Georgetown University Hospital/Medstar, Washington, District of Columbia; The Scripps Research Institute, La Jolla, California; and DNAX, Palo Alto, California

Chronic granulomatous disease (CGD) is a rare inherited disorderin which antimicrobial activity of phagocytes is impaired dueto the lack of reactive oxygen species, or oxidative burst,produced by NADPH oxidase. The X-linked form of CGD, representing $~$ 70% of all cases, is caused by mutations in the cytochrome bß subunit (CYBB) gene, which maps to chromosome Xp21.1.CYBB encodes the gp91-phox protein, a necessary component inthe NADPH oxidase pathway. A wide variety of mutations havebeen identified in X-linked CGD patients, all of which leadto deletion of the functional protein and no oxidative burstactivity. The mutations vary from single nucleotide substitutionsto deletions of the entire gene. In this article, we reporta mutation detection method for probands of female relativesat risk for carrier status of large deletions of the CYBB gene.Through fluorescent in situ hybridization of metaphase chromosomes,we were able to consistently distinguish carriers from noncarriersusing polymerase chain reaction-derived, labeled DNA specificfor exons 2 to 13 of the CYBB region at Xp21.1.