Difference between revisions of "Part:BBa K4201018:Design"
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Part information and design consideration can be found on respective basic parts pages. | Part information and design consideration can be found on respective basic parts pages. | ||
− | This construct differs from similar composite parts like | + | This construct differs from similar composite parts like BBa_K4201016 by having different promoter (Gmubi) and terminator (AtHSP) amounts and locations. In these constructs, dashes(-) represent locations where genes were synthesized <i>de novo</i>. An underscore (_) symbolizes a location where we adhered genes together via Golden Gate assembly. Utilizing Golden Gate assembly allowed us to insert promoter and terminator sequences between genes of interest, and consequently enables the study of transcription efficacy based on promoter and terminator location. |
+ | |||
+ | RUBY is a reporter gene that enables the visualization of transformation by changing the color of the plant from green to purle-red<sup>4</sup>. | ||
+ | |||
+ | Gmubi is constitutive promoter native to <i>Glyine max</i><sup>5</sup>, while AtHSP is the terminator of a heat shock protein that has shown to promote expression in plants<sup>6</sup>. | ||
+ | |||
+ | This assembly was made using NEB 10-<i>beta</i> cells and GoldBio EHA105 <i>agrobacterium</i> as intermediate hosts. Parts BBa_K4201013 (Amp BsaI) and BBa_K4201014 (Chlor AaRI) were used as intermediate backbones for Golden Gate assembly. The construct was integrated into part BBa_K4201015 (Kan BsaI) as a final backbone before transfection into <i>Glycine max</i>. | ||
===Source=== | ===Source=== | ||
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cytoTDS-MBP is a de novo synthesized construct made for this project. Learn more about our construct and its genes by searching for BBa_K4201006. | cytoTDS-MBP is a de novo synthesized construct made for this project. Learn more about our construct and its genes by searching for BBa_K4201006. | ||
− | RUBY is a reporter gene from the order Caryophyllales. | + | RUBY is a reporter gene from the order Caryophyllales<sup>4</sup>. |
− | Gmubi promoters originate from <i>Glycine max</i>. | + | Gmubi promoters originate from <i>Glycine max</i><sup>5</sup>. |
− | AtHSP terminators originate from the <i>Arabidopsis thaliana</i> genome. | + | AtHSP terminators originate from the <i>Arabidopsis thaliana</i> genome<sup>6</sup>. |
− | All genes were synthesized <i>de novo</i> by iGEM sponsors Twist Biosciences and IDT. | + | All genes were synthesized <i>de novo</i> by iGEM sponsors Twist Biosciences and IDT. |
===References=== | ===References=== | ||
+ | |||
+ | 1. Majer, E., Llorente, B., Rodríguez-Concepción, M. & Daròs, J.-A. Rewiring carotenoid biosynthesis in plants using a viral vector. Sci. Rep. 7, 41645 (2017). | ||
+ | |||
+ | 2. Xiong, X. et al. The Taxus genome provides insights into paclitaxel biosynthesis. Nat. Plants 7, 1026–1036 (2021). | ||
+ | |||
+ | 3. Lebendiker, M. & Danieli, T. Purification of Proteins Fused to Maltose-Binding Protein. Methods Mol. Biol. Clifton NJ 1485, 257–273 (2017). | ||
+ | |||
+ | 4. He, Y., Zhang, T., Sun, H., Zhan, H. & Zhao, Y. A reporter for noninvasively monitoring gene expression and plant transformation. Hortic. Res. 7, 1–6 (2020). | ||
+ | |||
+ | 5. De La Torre, C. M. & Finer, J. J. The intron and 5’ distal region of the soybean Gmubi promoter contribute to very high levels of gene expression in transiently and stably transformed tissues. Plant Cell Rep. 34, 111–120 (2015). | ||
+ | |||
+ | 6. Nagaya, S., Kawamura, K., Shinmyo, A. & Kato, K. The HSP Terminator of Arabidopsis thaliana Increases Gene Expression in Plant Cells. Plant Cell Physiol. 51, 328–32 (2010). |
Latest revision as of 10:23, 12 October 2022
CrtE_cytoTDS2-MBP_RUBY
- 10INCOMPATIBLE WITH RFC[10]Illegal XbaI site found at 4087
Illegal PstI site found at 2087
Illegal PstI site found at 3774
Illegal PstI site found at 5335
Illegal PstI site found at 6893
Illegal PstI site found at 12101 - 12INCOMPATIBLE WITH RFC[12]Illegal PstI site found at 2087
Illegal PstI site found at 3774
Illegal PstI site found at 5335
Illegal PstI site found at 6893
Illegal PstI site found at 12101 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 790
Illegal BglII site found at 1153
Illegal BglII site found at 2021
Illegal BglII site found at 2956
Illegal BglII site found at 6827
Illegal BglII site found at 7762
Illegal BglII site found at 11664
Illegal BamHI site found at 10313 - 23INCOMPATIBLE WITH RFC[23]Illegal XbaI site found at 4087
Illegal PstI site found at 2087
Illegal PstI site found at 3774
Illegal PstI site found at 5335
Illegal PstI site found at 6893
Illegal PstI site found at 12101 - 25INCOMPATIBLE WITH RFC[25]Illegal XbaI site found at 4087
Illegal PstI site found at 2087
Illegal PstI site found at 3774
Illegal PstI site found at 5335
Illegal PstI site found at 6893
Illegal PstI site found at 12101
Illegal NgoMIV site found at 1697
Illegal NgoMIV site found at 8249
Illegal NgoMIV site found at 8828
Illegal NgoMIV site found at 10541
Illegal AgeI site found at 3923 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 1
Illegal BsaI site found at 948
Illegal BsaI site found at 1887
Illegal BsaI site found at 2167
Illegal BsaI site found at 3114
Illegal BsaI site found at 6693
Illegal BsaI site found at 6973
Illegal BsaI.rc site found at 934
Illegal BsaI.rc site found at 1873
Illegal BsaI.rc site found at 2153
Illegal BsaI.rc site found at 3100
Illegal BsaI.rc site found at 6679
Illegal BsaI.rc site found at 6959
Illegal BsaI.rc site found at 7906
Design Notes
Part information and design consideration can be found on respective basic parts pages.
This construct differs from similar composite parts like BBa_K4201016 by having different promoter (Gmubi) and terminator (AtHSP) amounts and locations. In these constructs, dashes(-) represent locations where genes were synthesized de novo. An underscore (_) symbolizes a location where we adhered genes together via Golden Gate assembly. Utilizing Golden Gate assembly allowed us to insert promoter and terminator sequences between genes of interest, and consequently enables the study of transcription efficacy based on promoter and terminator location.
RUBY is a reporter gene that enables the visualization of transformation by changing the color of the plant from green to purle-red4.
Gmubi is constitutive promoter native to Glyine max5, while AtHSP is the terminator of a heat shock protein that has shown to promote expression in plants6.
This assembly was made using NEB 10-beta cells and GoldBio EHA105 agrobacterium as intermediate hosts. Parts BBa_K4201013 (Amp BsaI) and BBa_K4201014 (Chlor AaRI) were used as intermediate backbones for Golden Gate assembly. The construct was integrated into part BBa_K4201015 (Kan BsaI) as a final backbone before transfection into Glycine max.
Source
cytoTDS-MBP is a de novo synthesized construct made for this project. Learn more about our construct and its genes by searching for BBa_K4201006.
RUBY is a reporter gene from the order Caryophyllales4.
Gmubi promoters originate from Glycine max5.
AtHSP terminators originate from the Arabidopsis thaliana genome6.
All genes were synthesized de novo by iGEM sponsors Twist Biosciences and IDT.
References
1. Majer, E., Llorente, B., Rodríguez-Concepción, M. & Daròs, J.-A. Rewiring carotenoid biosynthesis in plants using a viral vector. Sci. Rep. 7, 41645 (2017).
2. Xiong, X. et al. The Taxus genome provides insights into paclitaxel biosynthesis. Nat. Plants 7, 1026–1036 (2021).
3. Lebendiker, M. & Danieli, T. Purification of Proteins Fused to Maltose-Binding Protein. Methods Mol. Biol. Clifton NJ 1485, 257–273 (2017).
4. He, Y., Zhang, T., Sun, H., Zhan, H. & Zhao, Y. A reporter for noninvasively monitoring gene expression and plant transformation. Hortic. Res. 7, 1–6 (2020).
5. De La Torre, C. M. & Finer, J. J. The intron and 5’ distal region of the soybean Gmubi promoter contribute to very high levels of gene expression in transiently and stably transformed tissues. Plant Cell Rep. 34, 111–120 (2015).
6. Nagaya, S., Kawamura, K., Shinmyo, A. & Kato, K. The HSP Terminator of Arabidopsis thaliana Increases Gene Expression in Plant Cells. Plant Cell Physiol. 51, 328–32 (2010).