Difference between revisions of "Part:BBa K3278005"
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pDawn is a blue light control system which can start the expression of target gene under the inducement of blue light. More information in BBa_K1075044 added by iGEM13_Bonn. | pDawn is a blue light control system which can start the expression of target gene under the inducement of blue light. More information in BBa_K1075044 added by iGEM13_Bonn. | ||
− | However, expression leakage and delay are the two main problems of | + | However, expression leakage and delay are the two main problems of pDawn system. Leakage refers to the expression of target protein without light. According to our measurement, samples strictly kept away from light still had significant expression of GFP after being cultured for one night. Therefore, we carefully studied the gene circuits of pDawn system [2] [3] and hypothesized that enhancing the strength of the promoter of YF1 may enhance the repression of protein expression in the dark, thus solving the leakage problem. |
In this part, we use the tac promoter to substitute the lacIq promoter in front of YF1 and Fixk2 and compare its performance with the original pDawn-mEGFP component based on both the efficiency of mEGFP expression and the leakage level. To examine mEGFP expression dynamics, different illuminating duration was used in our experiment. | In this part, we use the tac promoter to substitute the lacIq promoter in front of YF1 and Fixk2 and compare its performance with the original pDawn-mEGFP component based on both the efficiency of mEGFP expression and the leakage level. To examine mEGFP expression dynamics, different illuminating duration was used in our experiment. | ||
− | + | pDawn with tac promoter is abbreviated as <b>tac</b> below. | |
[Go for the protocol of characterization:https://2019.igem.org/Team:ShanghaiTech_China/LightControl] | [Go for the protocol of characterization:https://2019.igem.org/Team:ShanghaiTech_China/LightControl] |
Revision as of 13:14, 20 October 2019
pDawn with tac promoter
pDawn is a blue light control system which can start the expression of target gene under the inducement of blue light. More information in BBa_K1075044 added by iGEM13_Bonn.
However, expression leakage and delay are the two main problems of pDawn system. Leakage refers to the expression of target protein without light. According to our measurement, samples strictly kept away from light still had significant expression of GFP after being cultured for one night. Therefore, we carefully studied the gene circuits of pDawn system [2] [3] and hypothesized that enhancing the strength of the promoter of YF1 may enhance the repression of protein expression in the dark, thus solving the leakage problem.
In this part, we use the tac promoter to substitute the lacIq promoter in front of YF1 and Fixk2 and compare its performance with the original pDawn-mEGFP component based on both the efficiency of mEGFP expression and the leakage level. To examine mEGFP expression dynamics, different illuminating duration was used in our experiment.
pDawn with tac promoter is abbreviated as tac below.
[Go for the protocol of characterization:https://2019.igem.org/Team:ShanghaiTech_China/LightControl]
To see more changes on pDawn, go to the websites below.
origin pDawn: https://parts.igem.org/Part:BBa_K1075044
pDawn with T7 promoter: https://parts.igem.org/Part:BBa_K3278001
pDawn with mtac promoter: https://parts.igem.org/Part:BBa_K3278002
pDawn with tac promoter: https://parts.igem.org/Part:BBa_K3278005
pDawn with YF2: https://parts.igem.org/Part:BBa_K3278006
References:
[1] From Dusk till Dawn: One-Plasmid Systems for Light-Regulated Gene Expression. Robert Ohlendorf1, Roee R. Vidavski, Avigdor Eldar, Keith Moffat, and Andreas Möglich. Journal of Molecular Biology, 2012, Vol.416, pp. 534-542.
[2] Design and Signaling Mechanism of Light-Regulated Histidine Kinases. Andreas Möglich, Rebecca A. Ayers and Keith Moffat. Biophysical Journal, 2009, Vol.96 (3), pp. 1433-1444.
[3] Oxygen-Regulated In Vitro Transcription of Rhizobium meliloti nifA and fixK Genes. JEAN-MARC REYRAT,' MICHEL DAVID,' CASIMIR BLONSKI,2 PIERRE BOISTARD,' AND JACQUES BATUT'*. Journal of Bacteriology, 1993, pp. 6867-6872.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 2161
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 46
Illegal NgoMIV site found at 178
Illegal NgoMIV site found at 272
Illegal NgoMIV site found at 565
Illegal NgoMIV site found at 1059
Illegal NgoMIV site found at 1077
Illegal NgoMIV site found at 1167
Illegal AgeI site found at 397
Illegal AgeI site found at 1525 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 1620
Illegal BsaI.rc site found at 508