A wide methodological approach to identify a large duplication in CFTR gene in a CF patient uncharacterised by sequencing analysis Lucy Costantino, Damiana.

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A wide methodological approach to identify a large duplication in CFTR gene in a CF patient uncharacterised by sequencing analysis Lucy Costantino, Damiana Rusconi, Laura Claut, Carla Colombo, Francesca Novara, Valentina Paracchini, Luigi Porcaro, Patrizia Capasso, Orsetta Zuffardi, Manuela Seia Journal of Cystic Fibrosis Volume 10, Issue 6, Pages 412-417 (December 2011) DOI: 10.1016/j.jcf.2011.06.007 Copyright © 2011 European Cystic Fibrosis Society. Terms and Conditions

Fig. 1 Analysis of the cDNA and genomic structure;A) Schematic representation of the duplicated region, where duplicated exons are referred to with 6b′, 7′, etc.B) CFTRdup6b-16 cDNA analysis. The 2166bp PCR product obtained using primers F8f (located on exon 15) and 14R (located on exon 14b) on a 1.5% agarose gel. L, ladder; lane 1: Cf parent with the duplication; lane 2: wild type sample. The ladder used is 50bp ladder (Invitrogen).C) Sequencing of the 2166bp fragment with the reverse primer B15R, located on exon 7. The image shows the junction between the beginning of the duplication (exon 6B) and the wild type exons 15–16. Journal of Cystic Fibrosis 2011 10, 412-417DOI: (10.1016/j.jcf.2011.06.007) Copyright © 2011 European Cystic Fibrosis Society. Terms and Conditions

Fig. 2 MLPA results analysed by Coffalyser software . The histogram shows the duplication of exons 6b–16. The ratio values including 0.75–1.3 indicate a wild type expression; ratio values lower than 0.75 suggest a deletion whilst values higher than 1.3 indicate a duplication. The probe 07-116.6 CFTR exon 10 (F508del) is normal because the patient is heterozygous for the F508del. Journal of Cystic Fibrosis 2011 10, 412-417DOI: (10.1016/j.jcf.2011.06.007) Copyright © 2011 European Cystic Fibrosis Society. Terms and Conditions

Fig. 3 Detection by genomic CGH array of the duplication of exons 6b–16 in the CFTR gene. The duplication involves about 70kb (proximally: last normal oligo 116,948,976Mb, first duplicated one 116,963,849Mb; distally: last duplicated oligo 117,030,994Mb, first normal one 117,039,020Mb) harbouring no less than 9 exons when considering the internal edges of the duplication. Journal of Cystic Fibrosis 2011 10, 412-417DOI: (10.1016/j.jcf.2011.06.007) Copyright © 2011 European Cystic Fibrosis Society. Terms and Conditions