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Evaluation
of a novel
luciferase reporter construct:
a
positive control plasmid for reporter gene assay
Tewin Tencomnao1*,
Varaporn Rakkhitawatthana2 and Kanya Sukhontasing2
1Center
for Excellence in Omics-Nano Medical Technology Project,
Department of Clinical Chemistry, Faculty of Allied Health
Sciences, Chulalongkorn University,
Bangkok 10330, Thailand.
2Undergraduate
Program in Medical Technology, Faculty of Allied Health
Sciences, Chulalongkorn University,
Bangkok 10330, Thailand.
*Corresponding author. E-mail:
tewin.t@chula.ac.th. Tel: (662) 218-1081 ext. 313. Fax: (662) 218-1082.
Abbreviations:
bp, base pair; CAT, chloramphenicol acetyltransferase;
cmv,
cytomegalovirus;
FBS, fetal bovine serum; GFP, green fluorescent
protein; PCR, polymerase chain reaction; RPMI, Rosewell Park Memorial Institute;
and SV40,
simian virus 40.
Accepted
2 May, 2008 |
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Reporter gene technology has been increasingly important in
the post-genomic era to explain human complexity and
diversity. The pGL3-Basic vector has been prevalently used
as a tool for analyzing cis-acting elements critical
for transcriptional mechanisms. In this work, we constructed
and evaluated the pGL3-Basic plasmid containing the
cytomegalovirus (CMV) enhancer/promoter aiming to establish
a positive control of pGL3-Basic vector. Using a human
melanoma cell line UACC-903 for transient transfection, the
novel luciferase reporter construct, pGL3-CMV, showed an
extremely high transcriptional activity approximately
4,260-fold greater than that of pGL3-Basic, indicating its
qualification as a positive control for luciferase reporter
gene assays.
Key
words:
Reporter gene plasmid, luciferase assay, cytomegalovirus
promoter/enhancer, human melanoma cell line. |
