TITLE:
Construction of Vector for Functional Analysis of the Intrinsically Bent DNA in the ACE3 Replicator from Drosophila melanogaster
AUTHORS:
Douglas Vinícius Bassalobre de Freitas, Fabrícia Gimenes, Adriana Fiorini, Fabiana dos Santos Rando, Quirino Alves de Lima Neto, Maria Aparecida Fernandez
KEYWORDS:
DNA Replication; Gene Amplification; ACE3; Intrinsically Bent DNA; Drosophila melanogaster
JOURNAL NAME:
Open Journal of Genetics,
Vol.4 No.1,
March
5,
2014
ABSTRACT: DNA replication is a crucial process for species
survival, nevertheless it is not clear which factors define origin selection in
multicellular eukaryotes. Developmental gene amplification systems, such as the
one described during ovarian follicles development in Drosophila melanogaster, are
useful tools for studying of DNA replication process in these organisms. We
previously described that the well characterized third chromosome amplified
domain of D. melanogater displays
three intrinsically bent DNA sites: b1,
localized at an amplification control element (ACE3), b2 and b3, both localized
at the preferential origin ori-β.
This proposal aimed to construct a Drosophila transformation vector, which contains a short deletion at the ACE3, in order to
reduce the intrinsically bent DNA site b1, and analyze the functional role of
this site in the gene amplification process. Through a series of cloning steps,
we obtained a Big Parent vector derivative, containing a deletion at the
positions 176-180 bp, inside the ACE3. The generation of a Drosophila transformation vector displays a reduced intrinsically bent DNA site in the third
chromosome amplified domain, it will allow the
analysis of the functional role of this curvature in developmental gene amplification, providing new insights on replication initiation in D. melanogaster and the function of
intrinsically bent DNA sites.