Difference between revisions of "Part:BBa K2440005"
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==Methods== | ==Methods== | ||
| − | SV40 promoter | + | To characterize the SV40 promoter, renilla luciferase was used as the reporter gene to quantify the transcriptional strength of promoter. To be specific, plasmids carrying sv40 promoter-renilla luciferase cassette was transfected into liver carcinoma cell line HepG2 by lipo3000 under standard proytocol. Cells were cultures under standard DMEM High glucose medium under 5% CO2 and 37 ℃ with 10% FBS. Cells were harvested 0, 6, 12 and 24h after transfection, and renilla luciferase activity was measured by Beyotime™ Dual Luciferase Reporter Gene Assay Kit. |
==Results== | ==Results== | ||
Latest revision as of 02:22, 22 October 2019
SV40 promoter
This is a promoter gene derived from SV40. It could initiate the transcription of downstream gene.
Usage and Biology
The promoter is a DNA region that initiates the transcription of a particular gene. The promoter is located upstream of the same strand at the transcription initiation site of a particular gene. 1
At pmirGLO Vector, SV40 early enhancer/promotor is located at 426–844 nucleotides, with a length of 419 nucleotides. Its role is to initiates the transcription of the hRluc-neo fusion protein coding region.
Sequence and Features
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Contribution:NUDT_CHINA, 2019
Summary: Since SV40 promoter has never been characterized in HepG2 cells, This year we added quantitative characterization data of the SV40 promoter.
Accurate characterization of promoter behavior is essential for the rational design of functional synthetic transcription networks such as logic gates and oscillators[1]. To characterize the SV40 promoter, renilla luciferase was used as the reporter gene to quantify the transcriptional strength.
Methods
To characterize the SV40 promoter, renilla luciferase was used as the reporter gene to quantify the transcriptional strength of promoter. To be specific, plasmids carrying sv40 promoter-renilla luciferase cassette was transfected into liver carcinoma cell line HepG2 by lipo3000 under standard proytocol. Cells were cultures under standard DMEM High glucose medium under 5% CO2 and 37 ℃ with 10% FBS. Cells were harvested 0, 6, 12 and 24h after transfection, and renilla luciferase activity was measured by Beyotime™ Dual Luciferase Reporter Gene Assay Kit.
Results
Results showed significant increase of luciferase level within the first 24 hours of expression.
Figure 1. Renilla luciferase activity of SV40-rLuc tranfected HepG2 cells. Error bar represents SD of at least 3 biological replicates.
Reference
[1] Gagniuc P, Ionescu-Tirgoviste C. (Apr 2013)."Gene promoters show chromosome-specificity and reveal chromosome territories in humans" BMC Genomics. 14:278. PMID: 23617842 PMCID: PMC3668249 DOI: 10.1186/1471-2164-14-278