PEDIATRICS Vol. 107 No. 2 February 2001, pp. 280-286

Comprehensive Mutation Screening in a Cystic Fibrosis Center

Received Feb 2, 2000; accepted Jun 12, 2000.

Jeffrey J. Wine^*, Eugene Kuo^*, Gregory Hurlock^*, and Richard B. Moss

From the ^* Cystic Fibrosis Research Laboratory, Stanford University, Stanford, California, and  Department of Pediatrics, Stanford University School of Medicine, Stanford, California.

Objectives and Background.  The identities of a cystic fibrosis (CF) patient's CFTR mutations can influence therapeutic strategies, but because >800 CFTR mutations exist, cost-effective, comprehensive screening requires a multistage approach. Single-strand conformation polymorphism and heteroduplex analysis (SSCP/HA) can be an important part of mutation detection, but must be calibrated within each laboratory. The sensitivity of a combined commercial-SSCP/HA approach to genotyping in a large, ethnically diverse US center CF population has not been established.

Study Design.  We screened all 27 CFTR exons in 10 human participants who had an unequivocal CF diagnosis including a positive sweat chloride test and at least 1 unknown allele after commercial testing for the 70 most common mutations by SSCP/HA. These participants were compared with 7 participants who had negative sweat tests but at least 1 other CF-like symptom meriting complete genotyping.

Results.  For the 10 CF participants, we detected 11 of 16 unknown alleles (69%) and all 4 of the known alleles (100%), for an overall rate of 75% inpatients not fully genotyped by conventional 70 mutation screen. For 7 participants with negative sweat tests, we confirmed 1 identified mutation in 14 alleles and detected 3 additional mutations. Mutations detected in both groups included 7 missense mutations (S13F, P67L, G98R, S492F, G970D, L1093P, N1303K) and 9 deletion, frameshift, nonsense or splicing mutations (R75X, G542X, F508, 451-4588 bp, 5T, 663T, exon 13 frameshift, 1261+1GA and 3272-26AG). Three of these mutations were novel (G970D, L1093P, and 451-4588 bp¹). Thirteen other changes were detected, including the novel changes 1812-3 ins T, 4096-278 ins T, 4096-265 ins TG, and 4096-180 TG.

Conclusion.  When combined with the 70 mutation Genzyme test, SSCP/HA analysis allows for detection of >95% of the mutations in an ethnically heterogeneous CF center population. We discuss 5 possible explanations that could account for the few remaining undetected mutations.  Key words:  CFTR, alleles, SSCP, heteroduplex analysis.

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