Latest revision as of 21:31, 1 October 2024

4xMREd promoter

Usage and Biology

To develop a cell-based heavy metal sensor, our research group generated a series of synthetic MTF-1-responsive promoter constructs and evaluated their efficacy. Since Wang and colleagues (2004) suggested a high affinity of MTF-1 towards MREd we synthesized a promoter sequence containing four MREd elements at the positioning of the MREs of the MREwt promoter (K5317003) to exclude possible disruption of the MTF-1 and MRE interaction.

Cloning

Theoretical Part Design

The sequence of the MREd site was obtained from Seguin and colleagues (1987), as well as the positioning and spacer sequences (Searle et al., 1985). The construct was synthesized.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

Characterization

The promoter was analyzed by composing a gene cassette where its placed upstream of the reporter gene EGFP to assess the promoter's functionality and metal-dependent efficiency based on the fluorescent signal. Please visit the K5317010 registry entry to view the results.

References

Searle, P. F., Stuart, G. W., & Palmiter, R. D. (1985). Building a metal-responsive promoter with synthetic regulatory elements. Molecular and cellular biology, 5(6), 1480–1489. https://doi.org/10.1128/mcb.5.6.1480-1489.1985

Seguin, C., & Hamer, D. H. (1987). Regulation in vitro of metallothionein gene binding factors. Science (New York, N.Y.), 235(4794), 1383–1387. https://doi.org/10.1126/science.3103216

Wang, Y., Lorenzi, I., Georgiev, O., & Schaffner, W. (2004). Metal-responsive transcription factor-1 (MTF-1) selects different types of metal response elements at low vs. high zinc concentration. Biological chemistry, 385(7), 623–632. https://doi.org/10.1515/BC.2004.077

@@ Line 18: / Line 18: @@
 =Characterization=
-The promoter was analyzed by composing a gene cassette where its placed upstream of the reporter gene EGFP to assess the promoters functionality and metal-dependent efficiency based on the fluorescent signal. Please visit the <span class="plainlinks">[https://parts.igem.org/Part:BBa_K5317010 K5317010]</span> registry entry to view the results.
+The promoter was analyzed by composing a gene cassette where its placed upstream of the reporter gene EGFP to assess the promoter's functionality and metal-dependent efficiency based on the fluorescent signal. Please visit the <span class="plainlinks">[https://parts.igem.org/Part:BBa_K5317010 K5317010]</span> registry entry to view the results.
 =References=