Nurul Wahida, Ab. Ghani
(2008)
Overexpression of multivalent
transcription factor, ctcf in hela cell
lines.
Other.
Universiti Sains Malaysia.
(Submitted)
Abstract
Transformation of pCI-CTCF plasmid was done after receiving this recombinant
construct from Dr. Elena M. Klenova, Gene Regulation Laboratory, University of Essex,
UK. The plasmid was propagated in E.coli DHS-a competence cell. Following
transformation, a transformant containing the recombinant plasmid was selected and
cultured overnight in LB Broth. The plasmid was then extracted using a plasmid extraction
kit QIAprep Spin Miniprep Kit (QIAGEN, Germany). Purified plasmid was further
analysed and characterized. The plasmid was then transfected into HeLa cells using
Calcium Phosphate method. Following transfection, the HeLa cells were lysed and the
supernatant of the lysed cells was analysed using Sodium Dodecyl Sulfate Polyacrylamide
Gel Electrophoresis (SDS-PAGE) followed by Western Blotting using anti CTCF
monoclonal antibody. There is a positive expression of exogenous CTCF detected in the
lysate compare to the normal HeLa cell lines control. However due to short period of time
for this project, transfection-efficiency was not properly estimated. This work established
conventional, cheap and reasonable transfection method for future functional assay works
in the lab.
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