DNA sequencing errors in molecular diagnostics of filamin myopathy.
BACKGROUND
Filamin myopathy is a neuromuscular disorder manifesting with predominantly limb-girdle muscle weakness and in many patients with diaphragm paralysis and cardiomyopathy, caused by mutations in the filamin C (FLNC) gene. Molecular diagnosis of filamin myopathy based on direct DNA sequencing of coding exons is compromised by the presence of a high homology pseudogene (pseFLNC) located approximately 53.6 kb downstream of the functional FLNC gene on chromosome 7q.


METHODS
Molecular cloning, RT-PCR and real-time PCR methods were used to detect sequence differences between the FLNC and pseFLNC that are implicated in known or potential molecular diagnostic errors. Overall, 50 patients with a phenotype resembling filamin myopathy have been screened for mutations in FLNC.


RESULTS
FLNC sequence inconsistencies caused by the interference from pseFLNC were identified and diagnostic errors involving, in particular, the detection of the most frequent disease-causing FLNC p.W2710X mutation resolved. Mismatches between the FLNC and pseFLNC sequences were tabulated for future use.


CONCLUSIONS
We devise a strategy that allows one to discern mutations occurring in the functional FLNC from those harbored in pseFLNC, thus preventing possible complications in future research and patient genetic testing.
Odgerel
Zagaa
Z
National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892-9404, USA.
van der Ven
Peter F M
PF
Fürst
Dieter O
DO
Goldfarb
Lev G
LG
D001483:Base Sequence; D002897:Chromosomes, Human, Pair 7 / Q000235:genetics; D003001:Cloning, Molecular; D003285:Contractile Proteins / Q000235:genetics*; D004247:DNA / Q000235:genetics*; D005091:Exons; D064448:Filamins; D006801:Humans; D008840:Microfilament Proteins / Q000235:genetics*; D025202:Molecular Diagnostic Techniques; D009135:Muscular Diseases / Q000175:diagnosis* / Q000235:genetics*; D009154:Mutation; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA
D003285:Contractile Proteins; C578661:FLNC protein, human; D064448:Filamins; D008840:Microfilament Proteins; D004247:DNA