The diagnostics of myotonic dystrophy

Mendelovo Centrum pro vzdělání v biologii, biomedicíně a bioinformatice
a Centrum molekulární biologie a genové terapie FN
zvou na přednášku
The diagnostics of myotonic dystrophy
8.února 2010, v 14:30 hodin
posluchárna Centra molekulární biologie a genové terapie IHOK FN Brno
(pracoviště Dětská nemocnice, Černopolní 9, 613 00 Brno)
Mgr. Soňa Čejková
Abstract:
Myotonic dystrophy (DM) is an autosomal dominant disorder, its incidence is 1:8000. It is
multisystematic disorder including myotonia, muscle wasting, insulin resistance, cataract, atc. Two
types of DM exist: DM type one has trinucleotide (CTG)n expansion within DMPK 3´-untranslated
region, dystrophya myotonica protein kinase gene is localized on 19q, while DM type two has
tetranucleotide (CCTG)n expansion within intron 1 of the ZNF9, zinc finger 9 gene is localized on 3q.
How is DM diagnosed in our laboratory? We use fragmentation analysis or Southern blotting.
We use two combinations of PCR for fragmentation analysis. In the first combination of PCR we use
two primers flanking the gene and three primers in the second combination of PCR (P1 - is bound
before coding sequence; primer P4 - first part of this primer is bound to repeats and the second part
of this primer is a sequence, which is not in human genome; and primer P3 - complementary to the
second part of the primer P4). Products of PCRs are visualized by fragmentation analysis.
Southern blotting: we cut genomic DNA by EcoRI, after that we separate DNA fragments by agarose
gel electrophoresis. Then we continue with blotting of fragments to nitrocellulose membrane and
hybridization with DNA probe labelled by isotope of phosphor. At the end the fragments are
visualized by autoradiography.
Kontakt: [email protected]