Naslov
Validation of the Screening Method for Dynamic Mutations in the FMR1 Gene.

Autori
Škrlec, Ivana ; Tomac, Višnja ; Pušeljić, Silvija ; Galić, Mia ; Barišić, Karmela ; Wagner, Jasenka

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
10th ISABS Conference on Forensic and Anthropologic Genetics and Mayo Clinic Lectures in Individualized Medicine / - , 2017

ISBN
978-953-57695-2-1

Skup
10th ISABS Conference on Forensic and Anthropologic Genetics and Mayo Clinic Lectures in Individualized Medicine

Mjesto i datum
Dubrovnik, Hrvatska, 19.06.2017. - 23.06.2017

Vrsta sudjelovanja
Poster

Vrsta recenzije
Međunarodna recenzija

Ključne riječi
DNA denaturation ; fragile X syndrome ; intellectual disability ; FMR1 gene ; direct triplet-primed PCR method (dTP-PCR) ; CSHG

Sažetak
The objectives of this study are a validation of the direct triplet-primed PCR method (dTP-PCR) for determination of dynamic mutations in the FMR1 gene, and the comparison of the results of the dTP-PCR method and Southern blot analysis. In this study, 40 patients with a diagnosis of the intellectual disability were included. The patients were chosen randomly and the protocol was conducted as a blind study. The number of the CGG repeats in the FMR1 gene is determined by direct triplet-primed PCR method and melting curve analysis. Cut-off temperature between normal and permutation of the CGG repeats is determined by control samples with known number of CGG repeats. All patients are classified into four categories based on DNA melting curve (normal number of CGG repeats ≤44, gray zone 45 to 54 CGG repeats, premutation 55 to 200 CGG repeats, and full mutation >200 CGG repeats). A total of 10% patients had a full mutation (2.5% were female and 7.5% were men), and there were no patients with neither permutation nor mosaic samples. The presence of expanded alleles (>200 CGG repeats) in both sexes had different DNA melting curve than the melting curve of normal allele (<30 CGG repeat). The DNA melting curves of female heterozygote with expanded alleles differ from homozygous and heterozygous samples that contained normal alleles. The clinical performance of the assay was established with 40 previously analyzed samples, yielding results of 100% sensitivity and 90, 48% specificity in detection expansions of CGG (>30) repeats in FMR1 gene, while 89% of sensitivity and 99% specificity was achieved in the detection of expansion larger than 200 CGG repeats. The level of the accuracy of the dTP- PCR versus Southern blot method was 95%. This method is not suitable for determination of the methylation status. This method is appropriate for the quick determination of allelic changes in the FMR1 gene, screening population and defining mutations or premutation carriers in the population with intellectual disabilities with an unknown cause.

Izvorni jezik
Engleski

Znanstvena područja
Temeljne medicinske znanosti, Kliničke medicinske znanosti

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