[Structural characterization of 5' flanking regulatory region of DNA repair gene Rad51].
OBJECTIVE
To clarify the regulatory elements of Rad51 gene in its 5'flanking region.


METHODS
Various constructs were obtained by cloning different DNA fragments into pGL3 reporter vector. These constructs were then introduced into osteosarcoma cell line U2-OS by calcium phosphate method for transient expression of reporter gene, and luciferase activities were measured by luciferase assay.


RESULTS
Cells transfected with pGL3 constructs containing fragment -964 to +1430 and -733 to +1430 showed high luciferase activities. Obvious elevation of luciferase activities was also observed in cells transfected with pGL3 constructs containing four shorter derivative fragments -964 to -412, -746 to -412, -651 to -412 and -536 to -412. The highest luciferase activities were measured in transfected cells with plasmids containing fragment -964 to -412, and the lowest were in transfected cells with plasmids containing fragment -536 to -412. Luciferase activities in transfected cells with plasmids containing fragment -651 to -412 were higher than that in transfected cells with plasmids containing fragment -746 to -412.


CONCLUSIONS
It is believable that the basic transcription-promoting element (promoter) for Rad51 gene resides between -536 to -412, and two transcription-enhancing elements (enhancer) or binding sites of positive transcription factors reside between -651 to -536 and -964 to -746, whereas one transcription-inhibiting element (silencer) or binding site of negative transcription factor may reside between -746 to -651.
Yuan
Ying
Y
Cancer Institute, 2nd Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, 310009 PR China.
Ye
Jun
J
Dong
Qi
Q
Zheng
Shu
S
D024506:5' Flanking Region; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000235:genetics*; D006801:Humans; D011401:Promoter Regions, Genetic; D051135:Rad51 Recombinase
D004268:DNA-Binding Proteins; C495153:RAD51 protein, human; D051135:Rad51 Recombinase