Difference between revisions of "Part:BBa K346002:Design"
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In our design, ''merR'' was isolated from the operon and assembled with constitutive promoters of certain strength to maintain its expression intensity at certain level. For the same reason, the divergent promoter Pr was also removed by deletion of its -35 region. One biosensor construct was made by fusing PmerT and a reporting system, gfp, along with a plasmid structure that constitutive promoters prefixed before merR coding sequence. | In our design, ''merR'' was isolated from the operon and assembled with constitutive promoters of certain strength to maintain its expression intensity at certain level. For the same reason, the divergent promoter Pr was also removed by deletion of its -35 region. One biosensor construct was made by fusing PmerT and a reporting system, gfp, along with a plasmid structure that constitutive promoters prefixed before merR coding sequence. | ||
| + | [[Image:merT-merR.jpg]] | ||
===Source=== | ===Source=== | ||
Revision as of 01:35, 26 October 2010
PmerT promoter (mercury-responsive)
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
In our design, merR was isolated from the operon and assembled with constitutive promoters of certain strength to maintain its expression intensity at certain level. For the same reason, the divergent promoter Pr was also removed by deletion of its -35 region. One biosensor construct was made by fusing PmerT and a reporting system, gfp, along with a plasmid structure that constitutive promoters prefixed before merR coding sequence.
Source
PmerT is a promoter from Tn21 mercury resistance (mer) operon. By annealing PmerT-forward (5’-3’) and PmerT-reverse (5’-3’) primers, PmerT carrying a sticky end of EcoRI and SpeI was cloned into Psb1C3. The sequence is as below.
TTCCATATCGCTTGACTCCGTACATGAGTACGGAAGTAAGGTTACGCTATCCAATCC
References
Brown, N. L., J. V. Stoyanov, et al. (2003). "The MerR family of transcriptional regulators." FEMS Microbiol Rev 27(2-3): 145-163.
Hobman, J. L., J. Wilkie, et al. (2005). "A design for life: prokaryotic metal-binding MerR family regulators." Biometals 18(4): 429-436.
Liebert, C. A., R. M. Hall, et al. (1999). "Transposon Tn21, flagship of the floating genome." Microbiol Mol Biol Rev 63(3): 507-522.
Nakaya, R., A. Nakamura, et al. (1960). "Resistance transfer agents in Shigella." Biochem Biophys Res Commun 3: 654-659.
Park, S. J., J. Wireman, et al. (1992). "Genetic analysis of the Tn21 mer operator-promoter." J Bacteriol 174(7): 2160-2171.