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| Identifier |
uch.biology.msc//2005apostolopoulou |
| Title |
Δημιουργία πλασμιδιακής κατασκευής για τη στοχευμένη απενεργοποίηση του γονιδίου Fra10Ac1 στο ποντίκι |
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Author
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Αποστολοπούλου, Δέσποινα
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| Abstract |
During our team's effort to functionally characterize human chromosome 10 genes which have been associated with clinical phenotypes, we studied FRA10AC1 gene, located at FRA10A fragile site (10q23.3-q24.2). This site belongs to rare folate sensitive fragile sites and is the most frequent among autosomal rare fragile sites. Recently, we found out that FRA10A expression is directly linked to the extension and hypermethylation of a trinucleotide CGG repeat lying at the 5 UTR of FRA10AC1 gene. FRA10A expression has been associated with mental retardation. This work contributes to the generation of a mouse model, in which one or both Fra10Ac1 alleles will be inactivated. This will allow the detailed description of partial or total FRA10AC1 loss of function phenotype. Fra10Ac1 is unique in mouse genome. Thus, its inactivation by homologous recombination will contribute to its functional analysis. This work aims at the screening of mouse strain 129SV genomic library for the isolation of genomic clones that will be used for the inactivation of the endogenous gene, the designing of the final target construct and that of the necessary intermediate constructs, as well as their construction. Three different radiolabeled probes from genomic or Fra10Ac1 cDNA were used for the library screening. Each screening was performed on ~3x105 clones. Totally, six different unique clones, extending from exon 1 to exon 11, were isolated. Their integrity control and their mapping on the genomic sequence were obtained either by restriction analysis or PCR. One of these clones, containing the first exons of the gene, was selected for the designing of the intermediate and the final constructs. Finally, four intermediate constructs were generated and tested by restriction analysis and DNA sequencing. The generation of Fra10Ac1-/- mouse model can compensate for the absence of FRA10A homozygotes in humans. The detailed phenotype analysis of Fra10Ac1-/- and Fra10Ac1+/- animals, which mimic the total or partial Fra10Ac1 protein loss, respectively, must include brain histology studies and behavioral tests, as well as examination for mental abnormalities. Therefore, this model will contribute to the correlation of FRA10AC1 function with the FRA10A phenotype.
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| Language |
Greek |
| Issue date |
2005-11-28 |
| Collection
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School/Department--School of Sciences and Engineering--Department of Biology--Post-graduate theses
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Type of Work--Post-graduate theses
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| Permanent Link |
https://elocus.lib.uoc.gr//dlib/4/1/3/metadata-dlib-2005apostolopoulou.tkl
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| Views |
237 |
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