<p align="justify">We proudly present the Marburg Collection, a novel golden-gate-based toolbox containing various parts that are compatible with the PhytoBrick system and MoClo. Compared to other bacterial toolboxes, the Marburg Collection shines with superior flexibility. We overcame the rigid paradigm of plasmid construction - thinking in fixed backbone and insert categories - by achieving complete <i> de novo </i> assembly of plasmids.
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36 connectors facilitate flexible cloning of multigene constructs and even allow for the inversion of individual transcription units. Additionally, our connectors function as insulators to avoid undesired crosstalk.
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The Marburg Collection contains 123 parts in total, including:
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inducible promoters, reporters, fluorescence and epitope tags, oris, resistance cassettes and genome engineering tools. To increase the value of the Marburg Collection, we additionally provide detailed experimental characterization for <i> V. natriegens </i> and a supportive software. We aspire availability of our toolbox for future iGEM teams to empower accelerated progression in their ambitious projects.</p>
[[File:T--Marburg--Cloning_Overview.png|800px|thumb|left|'''Figure 3''': <b> Hierarchical cloning is facilitated by subsequent Golden Gate reactions. </b> <br> Basic building blocks like promoters or terminators are stored in level 0 plasmids. Parts from each category of our collection can be chosen to built level 1 plasmids harboring a single transcription unit. Up to five transcription units can be assembled into a level 2 plasmid.]]
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[[File:T--Marburg--Overview_Marburg_Toolbox.png|750px|thumb|middle|'''Figure 4''': <b> Additional bases and fusion sites ensure correct spacing and allow tags. </b> <br> Between some parts, additional base pairs were integrated to ensure correct spacing and to maintain the triplet code. We expanded our toolbox by providing N- and C- terminal tags by creating novel fusions and splitting the CDS and terminator part, respectively.]]
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Latest revision as of 16:30, 13 October 2022
Plac+TE2
consitutive promoter in bacteria and yeast
Sequence and Features
Assembly Compatibility:
10
INCOMPATIBLE WITH RFC[10]
Illegal XbaI site found at 279
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
INCOMPATIBLE WITH RFC[23]
Illegal XbaI site found at 279
25
INCOMPATIBLE WITH RFC[25]
Illegal XbaI site found at 279
1000
COMPATIBLE WITH RFC[1000]
BNSC-China
We crossed the constitutive promoters of the J23 family (J23119, J23100, J23104, J23113) in prokaryotes as well as plac with several artificially designed promoters (TE1, TE2, TE3, TE4, TE5) in yeast that have been recently reported in the literature. The bacterial constitutive promoter is loaded upstream of the yeast promoter to form a hybrid promoter. We tested these combinations in brewer's yeast as well as bacteria and found that these promoters stably drove gene expression in both yeast and bacteria, and showed a decreasing trend consistent with data reported in the literature. These data suggest that our design worked, and although these synthetic promoters may differ in intensity from when they work alone, at the functional level this can be left out of the discussion.
Figure. 7 Characterization of PVan_CC and PVan_CC_THS.
