A Frequent Large Rearrangement in theCFTRGene in Cystic Fibrosis Patients from Reunion Island

Abstract

Reunion Island is a French province, 800 km east of Madagascar and 200 km west of Mauritius. On ReunionIsland, the birth prevalence of cystic fibrosis (CF) is particularly high in the population of European origin, approximately 1:1000. In a previous study, we demonstrated that the screening of the 27 exons of the CF transmembrane conductance regulator (CFTR) gene by denaturing high-pressure liquid chromatography (DHPLC) in 114 CF families allowed the detection of about 93% of the molecular defects present on Reunion Island. Unidentified CF mutations may lie in introns or in regulatory regions that are not routinely investigated, or may correspond to gene rearrangements such as large, heterozygous deletions that escape detection using current PCR-based techniques. Using a combination of different methods (such as multiplex ligationdependent probe amplification), 6 of the 13 unidentified CF alleles (46%) were found to harbor a deletion of 5288 bp, spanning from exon 17a to 18. Identification and examination of the breakpoint sequences showed that this deletion is different from the 3120؉1kbdel8.6Kb previously found in the Palestinian Arabs. The chromosomes bearing IVS16؉3316_IVS18؉644del5288 did not have a common extragenic haplotype. Clinical evaluation of homozygotes (2 unrelated patients) and compound heterozygotes indicated that this deletion represents a severe mutation associated with positive sweat chloride test, pancreatic insufficiency, and early age at diagnosis. 208

Validation of a Custom Next-Generation Sequencing Assay for Cystic Fibrosis Newborn Screening

“…In addition to full gene sequence analysis, algorithms are used to impute large del/dup typically not detectable by sequencing. Most del/dup are individually rare [ 29 , 30 ], although several recurrent deletions with known breakpoints have been reported in CF patients [ 31 , 32 , 33 , 34 ] and have been detected in our population ( Table S8 ). CFTR del/dup are not routinely screened due to their rarity, and most are not detected using NGS panels because they are larger than can be consistently called using standard SNV/indel callers.…”

Multiplex Ligation-Dependent Probe Amplification Identification of Whole Exon and Single Nucleotide Deletions in the CFTR Gene of Hispanic Individuals with Cystic Fibrosis

“…However, with the advent of techniques such as quantitative multiplex PCR of short fluorescent fragments, semiquantitative fluorescent PCR, semiquantitative fluorescent multiplex PCR, and MLPA, more genomic rearrangements in the CFTR gene have recently been identified, suggesting that exon deletions and duplications are of clinical importance in CF etiology (Table 1). 17,19,[31][32][33][34][35][36][37][38][39][40][41][42][43][44][45][46][47][48] Our work is the first exon rearrangement study to specifically target Hispanic patients with CF.…”

Detection of two Alu insertions in the CFTR gene

“…sickkids.on.ca/cftr/app; as of December 6, 2006), N97% are either single base-pair substitutions or microinsertions/deletions. Recently, however, an increasing number of large genomic rearrangements have been reported in the CFTR gene, a development potentiated by the introduction of quantitative PCR-based techniques [5][6][7][8][9][10][11][12][13][14][15]. Of the ∼ 30 such mutational events already documented in the literature, 21 have been fully characterized at the nucleotide level [10].…”