Part:BBa_K5088675
Tarakate - Test construct GFP
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- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 230
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Background
For initial testing of our measurement system we used the GFP from the plant collection in the iGEM distribution kit. The expression was driven by the 35S CaMV promoter, TMV 5'UTR, and 35S 3'UTR. This system is widely used for constitutive overexpression of transgenes in various plant species, including model organisms and economically important crops (1).
Results
Figure 1: SBOL scheme of Tarakate - Test construct GFP [BBa_K5088675].
Figure 2: WT N. benthamiana plants treated with A. rhizogenes K599 and A. tumfaciens GV3101 P19 (negative control) under A) normal light conditions and B) under long-wave UV light (395nm). N. benthamiana infiltrated with Tarakate - Test construct GFP [BBa_K5088675] under C) normal light conditions and D) under long-wave UV light (395nm).
Figure 3: Successful expression of avGFP in N. benthamiana leaf disk assay using Tarakate - Test construct GFP [BBa_K5088675].
Figure 4: Successful expression of avGFP in protoplasts of Taraxacum officinale using Tarakate - Test construct GFP [BBa_K5088675].
Figure 5: Successful expression of avGFP in roots of T. kok-saghyz using the CDB protocol and Tarakate - Test construct GFP [BBa_K5088675]. A) Brightfield view of a transformed root of T. kok-saghyz B) The same root imaged under UV light.
The Dandelion Toolbox
Our project aimed to advance the genetic engineering of dandelions by developing a robust set of constitutive regulatory parts. Using a transcriptomic approach, we identified 40 endogenous elements. To ensure precise and reliable testing, we constructed a ratiometric measurement system, enabling effective and quantitative characterization of these parts.
We employed three distinct plant transformation methods to test and validate the functionality of the regulatory elements. Through rigorous testing, we successfully characterized 23 out of the initial 40 elements, resulting in a comprehensive collection of standardized dandelion parts. This well-characterized suite of parts is designed to streamline future complex genetic engineering projects.
By providing these standardized tools, our project significantly lowers the barriers for researchers and iGEM teams, making Taraxacum kok-saghyz a more accessible and versatile chassis for plant synthetic biology. Ultimately, our work contributes to enhancing dandelion as a model organism and supporting sustainable natural rubber production.
Overview
| Part Identifier | Part Type | Nickname | Part Description |
|---|---|---|---|
| BBa_K5088001 | Promoter + 5'UTR | P_RPL28 | Large subunit ribosomal protein L28e - Promoter+5'UTR from T. kok-saghyz |
| BBa_K5088006 | Promoter + 5'UTR | P_FKBP4_5 | FK506-binding protein 4/5 - Promoter+5'UTR from T. kok-saghyz |
| BBa_K5088007 | Promoter + 5'UTR | P_CLTC | Clathrin - Promoter+5'UTR from T. kok-saghyz |
| BBa_K5088008 | Promoter + 5'UTR | P_RPL31 | Large subunit ribosomal protein L31e - Promoter+5'UTR from T. kok-saghyz |
| BBa_K5088012 | Promoter + 5'UTR | P_Tubulin | Tubulin - Promoter+5'UTR from T. kok-saghyz |
| BBa_K5088013 | Promoter + 5'UTR | P_EIF5A | Translation initiation factor 5A - Promoter+5'UTR from T. kok-saghyz |
| BBa_K5088102 | 3'UTR | T_PTI1 | Protein tyrosine kinase - 3'UTR from T. kok-saghyz |
| BBa_K5088103 | 3'UTR | T_RPL28 | Large subunit ribosomal protein L28e - 3'UTR from T. kok-saghyz |
| BBa_K5088104 | 3'UTR | T_EPS15 | Epidermal growth factor receptor substrate 15 - 3'UTR from T. kok-saghyz |
| BBa_K5088105 | 3'UTR | T_GSK3B | Glycogen synthase kinase 3 - 3'UTR from T. kok-saghyz |
| BBa_K5088106 | 3'UTR | T_MGRN1 | E3 ubiquitin-protein ligase - 3'UTR from T. kok-saghyz |
| BBa_K5088107 | 3'UTR | T_RPL35A | Large subunit ribosomal protein L35Ae - 3'UTR from T. kok-saghyz |
| BBa_K5088108 | 3'UTR | T_betB | Betaine-aldehyde dehydrogenase - 3'UTR from T. kok-saghyz |
| BBa_K5088109 | 3'UTR | T_pgm | Phosphoglucomutase - 3'UTR from T. kok-saghyz |
| BBa_K5088110 | 3'UTR | T_ATP-synt | ATPase subunit gamma - 3'UTR from T. kok-saghyz |
| BBa_K5088111 | 3'UTR | T_EIF3B | Translation initiation factor 3 subunit B - 3'UTR from T. kok-saghyz |
| BBa_K5088112 | 3'UTR | T_RPL31 | Large subunit ribosomal protein L31e - 3'UTR from T. kok-saghyz |
| BBa_K5088113 | 3'UTR | T_TM9SF2_4 | Transmembrane 9 superfamily member 2/4 - 3'UTR from T. kok-saghyz |
| BBa_K5088114 | 3'UTR | T_CUL1 | Cullin - 3'UTR from T. kok-saghyz |
| BBa_K5088115 | 3'UTR | T_PSMB6 | 20S proteasome subunit beta 1 - 3'UTR from T. kok-saghyz |
| BBa_K5088116 | 3'UTR | T_RPSA | Small subunit ribosomal protein SAe - 3'UTR from T. kok-saghyz |
| BBa_K5088117 | 3'UTR | T_VPS4 | Vacuolar protein-sorting-associated protein 4 - 3'UTR from T. kok-saghyz |
| BBa_K5088118 | 3'UTR | T_EIF2S3 | Translation initiation factor 2 subunit 3 - 3'UTR from T. kok-saghyz |
Dandelion Handbook
By creating a suite of genetic tools and transformation methods, and sharing them through our Dandelion Handbook, we believe that dandelions can serve as an excellent chassis for numerous applications. We aim to inspire future iGEM teams to harness the unique properties of dandelions for a variety of promising projects.
Dandelions have demonstrated their versatility, being used as a coffee alternative and in various food applications such as salads, wine, and honey. Additionally, their ability to naturally hyperaccumulate environmental pollutants, including heavy metals, highlights their potential for bioremediation applications.
By equipping future iGEM teams with these resources, we aspire to unlock the full potential of dandelions, paving the way for sustainable and diverse synthetic biology applications.
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