Huntington's Disease is an inherited neurodegenerative disorder affecting human and caused by the expansion of a CAG repeat microsatellite localized in the 5’ end of the Huntingtin (Htt) gene, ultimately leading to neuronal loss in brain and motor and neuropsychiatric disturbances. To obtain information on Htt orthologous genes, a potential source of genomic DNA is represented by museum collections. Here we aimed at the characterization of orthologous sequences of the human microsatellite by assessing 141 specimens from Museo di Storia Naturale di Milano, Museo di Storia Naturale di Pavia and Acquario Civico di Milano. Samples belonged to four vertebrate groups: mammals (n=66), amphibians (n=1), fishes (n=22) and reptiles (n=52). Small fragments of tissues preserved in ethanol (n=52), stored frozen (n=79) and tissues that had undergone a tanning process (n=10) were used. These specimens ranged from recently collected to over 50-years-old specimens. Genomic DNA was extracted with two different commercial kits (Macherey-Nagel, Zymo Research) and its quality assessed by ultraviolet-visible spectrophotometry and gel electrophoresis. DNA extraction yielded fragments of highly variable length, probably depending on initial sample preservation conditions. Specific primers designed to encompass the CAG tract and targeting the most conservative regions of the different templates were used to amplify the sequences of interest. Amplicon length ranged from 100 to 250 base pairs. About 50% of the PCR reactions performed were successful and the relative products have been correctly sequenced. This study demonstrates that museum collections are a valuable resource of genomic material to amplify triplet repeat loci and restates the importance of specimens conservation for scientific research.
Successful sequencing of Huntington’s disease CAG microsatellite orthologs from museum collection specimens / G. Formenti, M. Pacifico, R. Iennaco, N. Ancona, G. Bardelli, M. Podestà, E. Razzetti, N. Saino, C. Zuccato, E. Cattaneo. ((Intervento presentato al 6. convegno SIBE tenutosi a Trento nel 2015.
|Titolo:||Successful sequencing of Huntington’s disease CAG microsatellite orthologs from museum collection specimens|
FORMENTI, GIULIO PAOLO (Primo)
CATTANEO, ELENA (Ultimo)
|Data di pubblicazione:||31-ago-2015|
|Parole Chiave:||Huntington's disease; CAG; repeat; evolution|
|Settore Scientifico Disciplinare:||Settore BIO/11 - Biologia Molecolare|
Settore BIO/18 - Genetica
|Citazione:||Successful sequencing of Huntington’s disease CAG microsatellite orthologs from museum collection specimens / G. Formenti, M. Pacifico, R. Iennaco, N. Ancona, G. Bardelli, M. Podestà, E. Razzetti, N. Saino, C. Zuccato, E. Cattaneo. ((Intervento presentato al 6. convegno SIBE tenutosi a Trento nel 2015.|
|Appare nelle tipologie:||14 - Intervento a convegno non pubblicato|