1
SUPPLEMENTARY INFORMATION
wt and 2657+5G>A mutant CFTR splicing minigene sequence
CTTTGGCATTAGGAGCTTGAGCCCAGACGGCCCTAGCAGGGACCCCAGCGCCCGAGAGACCATGCAGAGGTCGCCTCT
GGAAAAGGCCAGCGTTGTCTCCAAACTTTTTTTCAGCTGGACCAGACCAATTTTGAGGAAAGGATACAGACAGCGCCT
GGAATTGTCAGACATATACCAAATCCCTTCTGTTGATTCTGCTGACAATCTATCTGAAAAATTGGAAAGAGAATGGGA
TAGAGAGCTGGCTTCAAAGAAAAATCCTAAACTCATTAATGCCCTTCGGCGATGTTTTTTCTGGAGATTTATGTTCTA
TGGAATCTTTTTATATTTAGGGGAAGTCACCAAAGCAGTACAGCCTCTCTTACTGGGAAGAATCATAGCTTCCTATGA
CCCGGATAACAAGGAGGAACGCTCTATCGCGATTTATCTAGGCATAGGCTTATGCCTTCTCTTTATTGTGAGGACACT
GCTCCTACACCCAGCCATTTTTGGCCTTCATCACATTGGAATGCAGATGAGAATAGCTATGTTTAGTTTGATTTATAA
GAAGACTTTAAAGCTGTCAAGCCGTGTTCTAGATAAAATAAGTATTGGACAACTTGTTAGTCTCCTTTCCAACAACCT
GAACAAATTTGATGAAGGACTTGCATTGGCACATTTCGTGTGGATCGCTCCTTTGCAAGTGGCACTCCTCATGGGGCT
AATCTGGGAGTTGTTACAGGCGTCTGCCTTCTGTGGACTTGGTTTCCTGATAGTCCTTGCCCTTTTTCAGGCTGGGCT
AGGGAGAATGATGATGAAGTACAGAGATCAGAGAGCTGGGAAGATCAGTGAAAGACTTGTGATTACCTCAGAAATGAT
CGAGAACATCCAATCTGTTAAGGCATACTGCTGGGAAGAAGCAATGGAAAAAATGATTGAAAACTTAAGACAAACAGA
ACTGAAACTGACTCGGAAGGCAGCCTATGTGAGATACTTCAATAGCTCAGCCTTCTTCTTCTCAGGGTTCTTTGTGGT
GTTTTTATCTGTGCTTCCCTATGCACTAATCAAAGGAATCATCCTCCGGAAAATATTCACCACCATCTCATTCTGCAT
TGTTCTGCGCATGGCGGTCACTCGGCAATTTCCCTGGGCTGTACAAACATGGTATGACTCTCTTGGAGCAATAAACAA
AATACAGGATTTCTTACAAAAGCAAGAATATAAGACATTGGAATATAACTTAACGACTACAGAAGTAGTGATGGAGAA
TGTAACAGCCTTCTGGGAGGAGGGATTTGGGGAATTATTTGAGAAAGCAAAACAAAACAATAACAATAGAAAAACTTC
TAATGGTGATGACAGCCTCTTCTTCAGTAATTTCTCACTTCTTGGTACTCCTGTCCTGAAAGATATTAATTTCAAGAT
AGAAAGAGGACAGTTGTTGGCGGTTGCTGGATCCACTGGAGCAGGCAAGACTTCACTTCTAATGATGATTATGGGAGA
ACTGGAGCCTTCAGAGGGTAAAATTAAGCACAGTGGAAGAATTTCATTCTGTTCTCAGTTTTCCTGGATTATGCCTGG
CACCATTAAAGAAAATATCATCTTTGGTGTTTCCTATGATGAATATAGATACAGAAGCGTCATCAAAGCATGCCAACT
AGAAGAGGACATCTCCAAGTTTGCAGAGAAAGACAATATAGTTCTTGGAGAAGGTGGAATCACACTGAGTGGAGGTCA
ACGAGCAAGAATTTCTTTAGCAAGAGCAGTATACAAAGATGCTGATTTGTATTTATTAGACTCTCCTTTTGGATACCT
AGATGTTTTAACAGAAAAAGAAATATTTGAAAGCTGTGTCTGTAAACTGATGGCTAACAAAACTAGGATTTTGGTCAC
TTCTAAAATGGAACATTTAAAGAAAGCTGACAAAATATTAATTTTGAATGAAGGTAGCAGCTATTTTTATGGGACATT
TTCAGAACTCCAAAATCTACAGCCAGACTTTAGCTCAAAACTCATGGGATGTGATTCTTTCGACCAATTTAGTGCAGA
AAGAAGAAATTCAATCCTAACTGAGACCTTACACCGTTTCTCATTAGAAGGAGATGCTCCTGTCTCCTGGACAGAAAC
AAAAAAACAATCTTTTAAACAGACTGGAGAGTTTGGGGAAAAAAGGAAGAATTCTATTCTCAATCCAATCAACTCTAT
ACGAAAATTTTCCATTGTGCAAAAGACTCCCTTACAAATGAATGGCATCGAAGAGGATTCTGATGAGCCTTTAGAGAG
AAGGCTGTCCTTAGTACCAGATTCTGAGCAGGGAGAGGCGATACTGCCTCGCATCAGCGTGATCAGCACTGGCCCCAC
GCTTCAGGCACGAAGGAGGCAGTCTGTCCTGAACCTGATGACACACTCAGTTAACCAAGGTCAGAACATTCACCGAAA
GACAACAGCATCCACACGAAAAGTGTCACTGGCCCCTCAGGCAAACTTGACTGAACTGGATATATATTCAAGAAGGTT
ATCTCAAGAAACTGGCTTGGAAATAAGTGAAGAAATTAACGAAGAAGACTTAAAGgtaggtatacatcgcttgggggt
atttcaccccacagaatgcattgagtagaatgcaatatgtagcatgtaacaaaatttactaaaatcataggattagga
taaggtgtatcttaaaactcagaaagtatgaagttcattaattatacaagcaacgttaaaatgtaaaataacaaatga
tttctttttgcaatggacatatctcttcccataaaatgggaaaggatttagtttttggtcctctactaagccagtgat
aactgtgactataagttagaaagcatttgctttattaccatcttgaaccctctgtgggaagaggtgcagtataaataa
ctgtataaataaatagtagctttcattatttatagctcgcaaaataatctgtatggaagtagcatatataaggtatat
aaacatttagcctcttgataggactaactcacattctggtttgtatatcagtcttgcctgaatttagctagtgtgggc
ttttttttatcttgtgagtttgctttatacattgggtttctgaaaagatttcttttagagaatgtatataagcttaac
atgtactagtgccaatcttcagacagaaattttgttctattaggttttaagaataaaagcattttatttttaaaacag
gaaataatataaaaaggagagtttttgttgttttagtagaaaacttaatgccttggatgaaatgagccatgggcaggg
ttgtaatgaattgatatgtttaatagtatagatcatttgtgaataatatgacctttgacaagacacaagccattaaca
tctgtaggcagaagtttccttctttgtaaaatgagggaataaaatagatccctaaagtgtgtaattttagtatttcta
aactttatgaaggtttcctaaatgataattcatctatatagtgtttttttgtgtgtttgtttgtttgtttgtttgaga
tggagtctcgctctgtcacctaggctggagtgcaatggtgcaacctcggctcactgcaacctctgcctcctgggttca
agctaatctcctgcctcagcctcctgagtagctgagattacaggcatgcaccaccatgccgagctaatttttgtattt
ttagtagagaaggggtttcatcatgttgaccaggctggtcttgaactcctgaccttgtgatccacccacctcagcctc
ccaaagtgctggtattacaggcgtgtgccaccacgtccagcctgagccactgcgcccagcccatctatatagtttaat
atcaatctaaatgaatttctcagtcctgagcctaaaaatttagttgtaaagaatgatatccttgactaataatagttt
ctattaatggattgcatctagtgctaggtggcatatatttagtccccacaactaccctggaaggtatttaaaattttt
cacatttgcagataaggaaactaaagttcagagttcggcaacatgcttgaattcaagcagctcctaggatgttaatgg
tggaggttgggttcaaatccagatctgtctgactcaaaaaatgcatactcctaaccagtgcactatatcccaattcca
taggagcccttctttgtgattcatagcactttcccatgagttttgttgattttgtgagaaacaaaactctttttcctt
2
tggactgtctggaatctctcttttacaaatttttgaaatgtatttctatgccaaaagacaaagatttctagaggaata
tgcctaggatgagaattatgtaatttaaatcacagctggaaagagagaaagtcctaagttactaagaaatgttcaaac
acaaatgggctttcagtctattggaagacctttatagctagaagtatactgaactgtacttgtccatggacccctgaa
gaaacaggttaaatcaaagagagttctgggaaacttcatttagatggtatcattcatttgataaaaggtatgccactg
ttaagcctttaatggtaaaattgtccaataataatacagttatataatcagtgatacatttttagaattttgaaaaat
tacgatgtttctcatttttaataaagctgtgttgctccagtagacattattctggctatagaatgacatcatacatgg
catttataatgatttatatttgttaaaatacacttagattcaagtaatactattcttttattttcatatattaaaaat
aaaaccacaatggtggcatgaaactgtactgtcttattgtaatagccataattcttttattcagGAGTGCCTTTTTGA
TGATATGGAGAGCATACCAGCAGTGACTACATGGAACACATACCTTCGATATATTACTGTCCACAAGAGCTTAATTTT
TGTGCTAATTTGGTGCTTAGTAATTTTTCTGGCAGAGgtaagaatgttctattgtaaagtattactggatttaaagtt
aaattaagatagtttggggatgtatacatatatatgcacacacataaatatgtatatatacacatgtatacatgtata
agtatgcatatatacacacatatatcactatatgtatatatgtatatattacatatatttgtgattttacagtatata
atggtatagattcatatagttcttagcttctgaaaaatcaacaagtagaaccactactgatagccagcagtctgagat
tagtttctttaaaaaatgtttattatttaaaacattcagctgtgatcttggctttcttgtgaggttcaatagtttcta
ttgagtaaaggagagaaatggcagagaatttacttcagtgaaatttgaattccattaacttaatgtggtctcatcaca
aataatagtacttagaacacctagtacagctgctggacccaggaacacaaagcaaaggaagatgaaattgtgtgtacc
ttgatattggtacacacatcaaatggtgtgatgtgaatttagatgtgggcatgggaggaataggtgaagatgttagaa
aaaaaatcaactgtgtcttgttccattccagGTGGCTGCTTCTTTGGTTGTGCTGTGGCTCCTTGGAAAgtgagtatt
ccatgtcctattgtgtagattgtgttttatttctgttgattaaatattgtaatccactatgtttgtatgtattgtaat
ccactttgtttcatttctcccaagcattatggtagtggaaagataaggttttttgtttaaatgatgaccattagttgg
gtgaggtgacacattcctgtagtcctagctcctccacaggctgacgcaggaggatcacttgagcccaggagttcaggg
ctgtagtgttgtatcattgtgagtagccaccgcactccagcctggacaatatagtgagatcctatatctaaaataaaa
taaaataaaatgaataaattgtgagcatgtgcagctcctgcagtttctaaagaatatagttctgttcagtttctgtga
aacacaataaaaatatttgaaataacattacatatttagggttttcttcaaattttttaatttaataaagaacaactc
aatctctatcaatagtgagaaaacatatctattttcttgcaataatagtatgattttgaggttaagggtgcatgctct
tctaatgcaaaatattgtatttatttagactcaagtttagttccatttacatgtattggaaattcagtaagtaacttt
ggctgccaaataacgatttcctatttgctttacagCACTCCTCTTCAAGACAAAGGGAATAGTACTCATAGTAGAAAT
AACAGCTATGCAGTGATTATCACCAGCACCAGTTCGTATTATGTGTTTTACATTTACGTGGGAGTAGCCGACACTTTG
CTTGCTATGGGATTCTTCAGAGGTCTACCACTGGTGCATACTCTAATCACAGTGTCGAAAATTTTACACCACAAAATG
TTACATTCTGTTCTTCAAGCACCTATGTCAACCCTCAACACGTTGAAAGCAGGTGGGATTCTTAATAGATTCTCCAAA
GATATAGCAATTTTGGATGACCTTCTGCCTCTTACCATATTTGACTTCATCCAGTTGTTATTAATTGTGATTGGAGCT
ATAGCAGTTGTCGCAGTTTTACAACCCTACATCTTTGTTGCAACAGTGCCAGTGATAGTGGCTTTTATTATGTTGAGA
GCATATTTCCTCCAAACCTCACAGCAACTCAAACAACTGGAATCTGAAGGCAGGAGTCCAATTTTCACTCATCTTGTT
ACAAGCTTAAAAGGACTATGGACACTTCGTGCCTTCGGACGGCAGCCTTACTTTGAAACTCTGTTCCACAAAGCTCTG
AATTTACATACTGCCAACTGGTTCTTGTACCTGTCAACACTGCGCTGGTTCCAAATGAGAATAGAAATGATTTTTGTC
ATCTTCTTCATTGCTGTTACCTTCATTTCCATTTTAACAACAGGAGAAGGAGAAGGAAGAGTTGGTATTATCCTGACT
TTAGCCATGAATATCATGAGTACATTGCAGTGGGCTGTAAACTCCAGCATAGATGTGGATAGCTTGATGCGATCTGTG
AGCCGAGTCTTTAAGTTCATTGACATGCCAACAGAAGGTAAACCTACCAAGTCAACCAAACCATACAAGAATGGCCAA
CTCTCGAAAGTTATGATTATTGAGAATTCACACGTGAAGAAAGATGACATCTGGCCCTCAGGGGGCCAAATGACTGTC
AAAGATCTCACAGCAAAATACACAGAAGGTGGAAATGCCATATTAGAGAACATTTCCTTCTCAATAAGTCCTGGCCAG
AGGGTGGGCCTCTTGGGAAGAACTGGATCAGGGAAGAGTACTTTGTTATCAGCTTTTTTGAGACTACTGAACACTGAA
GGAGAAATCCAGATCGATGGTGTGTCTTGGGATTCAATAACTTTGCAACAGTGGAGGAAAGCCTTTGGAGTGATACCA
CAGAAAGTATTTATTTTTTCTGGAACATTTAGAAAAAACTTGGATCCCTATGAACAGTGGAGTGATCAAGAAATATGG
AAAGTTGCAGATGAGGTTGGGCTCAGATCTGTGATAGAACAGTTTCCTGGGAAGCTTGACTTTGTCCTTGTGGATGGG
GGCTGTGTCCTAAGCCATGGCCACAAGCAGTTGATGTGCTTGGCTAGATCTGTTCTCAGTAAGGCGAAGATCTTGCTG
CTTGATGAACCCAGTGCTCATTTGGATCCAGTAACATACCAAATAATTAGAAGAACTCTAAAACAAGCATTTGCTGAT
TGCACAGTAATTCTCTGTGAACACAGGATAGAAGCAATGCTGGAATGCCAACAATTTTTGGTCATAGAAGAGAACAAA
GTGCGGCAGTACGATTCCATCCAGAAACTGCTGAACGAGAGGAGCCTCTTCCGGCAAGCCATCAGCCCCTCCGACAGG
GTGAAGCTCTTTCCCCACCGGAACTCAAGCAAGTGCAAGTCTAAGCCCCAGATTGCTGCTCTGAAAGAGGAGACAGAA
GAAGAGGTGCAAGATACAAGGCTTTAGAGAGCAGCATAAATGTTGACATGGGACATTTGCTCATGGAATTGGAGCTCG
TGGGACAGTCACCTCATGGAATTGGAGCTCGTGGAACAGTTACCTCTGCCTCAGAAAACAAGGATGAATTAAGTTTTT
TTTTAAAAAAGAAACATTTGG
Supp. Figure S1 wild-type and 2657+5G>A mutant CFTR cDNA sequence plus IVS 14, IVS 15
and IVS 16. CFTR minigenes were produced by inserting the complete wt-CFTR cDNA and
relevant intronic regions into the pCDNA5/FRT/CFTR mammalian expression vector, as
described previously (Masvidal, et al, 2014). Thus, the full sequence of the minigenes contains
3
full-length CFTR cDNA, relevant intronic regions and part of the 3’UTR region. The ATGtranslation initiation codon and the termination codon are highlighted in bold. Exons are
represented in capital letters and introns (IVS14, IVS15 and IVS16) in small letters. Exon 16 is
highlighted in grey. The 2657+5G>A mutation is underlined in bold and grey. The 2657+5G>A
mutant minigene sequence is the same as the wt sequence, as the former was produced from the
pCDNA5/FRT/wt-CFTR backbone by site-directed mutagenesis.
2657+5G>A CFTR splicing minigene intronic region targeted by AONs and their sequence
3’ss
15
5’ss
16
+5
+10
IVS16
+28
GTGAGTATTCCATGTCCTATTGTGTAGATTGTGTTTT
GGUACAGGAUAACACAUCU
AGGUACAGGAUAACACAUC
AG
17
CFTR pre-mRNA
Anti-AON1
Anti-AON2
Supp. Figure S2 Antisense oligonucleotide design and diagram showing an inset of CFTR IVS
16, which contains the 2657+5G>A splicing mutation, highlighted in bold (position +5). The
region targeted by the antisense oligonucleotides AON1 (+10 to +28) and AON2 and their
aligned sequences are represented below. As the 2657+5G>A mutation is located at the 5’ end of
IVS 16, 5 nucleotides downstream the intron, a region that is conserved among mammals and
harbours the canonical base-pairing region for U1 snRNA, a key component of U1 snRNP
required for correct splicing, the AONs were designed a few nucleotides downstream from the
U1 snRNA binding region. Nucleotides involved in base pairing with U1 snRNA are depicted in
the grey box. AON2 was designed targeting the same sequence as AON1 but shifted by one
nucleotide upstream (+9 to +27). 5′ ss: splicing donor site, 3’ ss: splicing acceptor site.
4
AON1 effect on the 2562T>G and 2657+5G>A CFTR variants assessed by sqRT-PCR in HEK293 and CFBE cells
HEK293
CFBE
AON1
wt
% exon 16 100
inclusion
SNP
100
Mut
55
Double
Mut
43
SNP
100
Mut
94
AON1
Double
Mut
94
wt
100
SNP
100
Mut
75
Double
Mut
68
SNP
100
Mut
94
Double
Mut
14
15
16
14
15
17
14
17
17
93
Supp. Figure S3 AON1 effect on the 2562T>G and 2657+5G>A CFTR variants using splicing
minigenes. The c.2562T>G is a common single nucleotide polymorphism (SNP) in exon 15
which we have previously found in patient samples to be in linkage disequilibrium with the
c.2657+5G>A mutation (Mut) (Masvidal L, et al, 2014). HEK293 and CF bronchial epithelial
(CFBE41o-; from herein CFBE) cells were transiently transfected for 24 h with wt or mutant
CFTR splicing minigenes alone or cotransfected with AON1. The AON1 effect on the SNP, both
alone (SNP) and in combination with the 2657+5G>A splicing mutation (double Mut) was
assessed by semi-quantitative (sq) RT-PCR. Left panel, sqRT-PCR analysis of wt, SNP, Mut,
double Mut and the AON1 effect on these variants in HEK293 cells. The same for the right hand
panel but in CFBE cells. Result from agarose gel electrophoretic analysis where the upper band
corresponds to correctly spliced products and lower band corresponds to alternatively spliced
products, i.e. exon 16 skipping. A lower band lacking exon 15 and 16 is detected, as previously
(Masvidal L, et al, 2014). Numbers shown below correspond to the average percentage levels of
exon 16 inclusion.
5
wt and alternative transcripts after AON1 correction confirmed by PCR-band sequencing
wt transcript- upper band
Mut
AON1
Mut
AON1
Mut
AON1
GAGTGCTTTTTTGATGATATGGAGAGCATACCAGCAGTGACTACATGGAACACATACCTTCGATATATTACTGTCCACAAGAGC
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
GAGTGCTTTTTTGATGATATGGAGAGCATACCAGCAGTGACTACATGGAACACATACCTTCGATATATTACTGTCCACAAGAGC
TTAATTTTTGTGCTAATTTGGTGCTTAGTAATTTTTCTGGCAGAGGTGGCTGCTTCTTTGGTTGTGCTGTGGCTCCTTGGAAAC
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
TTAATTTTTGTGCTAATTTGGTGCTTAGTAATTTTTCTGGCAGAGGTGGCTGCTTCTTTGGTTGTGCTGTGGCTCCTTGGAAAC
ACTCCTCTTCAAGACAAAGGGAATAGTACTCATAGTAGAAATAAC
|||||||||||||||||||||||||||||||||||||||||||||
ACTCCTCTTCAAGACAAAGGGAATAGTACTCATAGTAGAAATAAC
Alternative transcript- lower band (no ex16)
Mut
AON1
Mut
AON1
Mut
AON1
GAGTGCTTTTTTGATGATATGGAGAGCATACCAGCAGTGACTACATGGAACACATACCTTCGATATATTACTGTCCACAAGAGC
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
GAGTGCTTTTTTGATGATATGGAGAGCATACCAGCAGTGACTACATGGAACACATACCTTCGATATATTACTGTCCACAAGAGC
TTAATTTTTGTGCTAATTTGGTGCTTAGTAATTTTTCTGGCAGAGCACTCCTCTTCAAGACAAAGGGAATAGTACTCATAGTAG
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
TTAATTTTTGTGCTAATTTGGTGCTTAGTAATTTTTCTGGCAGAGCACTCCTCTTCAAGACAAAGGGAATAGTACTCATAGTAG
AAATAACAGCTATGCAGTGATTATCACCAGCAC
|||||||||||||||||||||||||||||||||
AAATAACAGCTATGCAGTGATTATCACCAGCAC
Supp. Figure S4 Alignment of wt and alternatively spliced transcripts after AON1 correction.
PCR products from upper (wt) and lower (no exon 16) bands were PCR re-amplified and
sequenced to confirm the presence of complete exon 16 inclusion after AON1 treatment. Upper
panel shows the alignment of wt transcripts from upper mutant PCR band vs upper mutant-AON1
treated PCR band, showing exon 15, exon 16 (highlighted in bold) and part of exon 17. Lower
panel shows the alignment of alternatively spliced products from the lower mutant PCR band vs
lower mutant-AON1 treated PCR band showing exon 15 and part of exon 17 with exon 16
missing.
6
Alternative splicing analysis by sqRT-PCR after kinetin and EGCG treatment
Mut
DMSO
K
EGCG
14
15
16
14
15
17
17
Supp. Figure S5 Small-molecule compound treatment of HEK293 Flp-In cells stably expressing
the 2657+5G>A CFTR splicing minigene. RT-PCR analysis of mutant (Mut) minigene transcripts
treated for 24 h with dimethyl sulfoxide (DMSO) used as control, kinetin (K; 200 μM) and
epigallocatechin gallate (EGCG; 50 μM). Other compounds have also been tested (data not
shown). These compounds have been previously shown to improve exon inclusion in other
disorders (Hims, et al, 2007; Anderson, et al, 2003; Sumanasekera, et al, 2008).
Supp. Table S1 Primer sequences for mCherry insertion upstream of CFTR minigenes and Flagtag insertion on IVS 17.
Oligonucleotide
Sequence (5’-3’)
mCherry-Fusion-CFTR fw
CGGATCCACTAGTAACGGCCATGGTGAGCAAGGGCGAGGAG
mCherry-Fusion-CFTR rv
GGCCTTTTCCAGAGGCGACCTCTGCATGGTGGGCCCGCGGTACCGTCGACT
Flag-tag fw
AAAGGGAATAGTACTCATAGTAGAAATAACGATTACAAAGATGATGATGAT
AAAAGCTATGCAGTG
Flag-tag rv
CACTGCATAGCTTTTATCATCATCATCTTTGTAATCGTTATTTCTACTATGAG
TACTATTCCCTTT
References
Anderson SL, Qiu J, Rubin BY. 2003. EGCG corrects aberrant splicing of IKAP mRNA in cells
from patients with familial dysautonomia. Biochem Biophys Res Commun 310(2):62733.
Hims MM, Ibrahim EC, Leyne M, Mull J, Liu L, Lazaro C, Shetty RS, Gill S, Gusella JF, Reed
R, Slaugenhaupt SA. 2007. Therapeutic potential and mechanism of kinetin as a treatment
for the human splicing disease familial dysautonomia. J Mol Med 85(2):149-61.
Masvidal L, Igreja S, Ramos MD, Alvarez A, de Gracia J, Ramalho A, Amaral MD, Larriba S,
Casals T. 2014. Assessing the residual CFTR gene expression in human nasal epithelium
cells bearing CFTR splicing mutations causing cystic fibrosis. Eur J Hum Genet
22(6):784-91.
Sumanasekera C, Watt DS, Stamm S. 2008. Substances that can change alternative splice-site
selection. Biochem Soc Trans 36(Pt 3):483-90.