Our project introduces a modular collection of newly designed genetic parts assembled to enable fluorescence, secretion, uptake, metabolic production, and biosafety control. Using Golden Gate cloning into the broad-host-range backbone pJUMP24-1A, we constructed plasmids combining functional geneblocks and safety systems.
This modular approach allowed us to generate Level 2 assemblies encoding fluorescent reporters (GFP, mCherry), therapeutic modules (L-DOPA biosynthesis), and kill switches for biosafety. Importantly, the design is fully interchangeable: future iGEM teams can replace or recombine parts to adapt the system for new functions or host organisms.
Taken together, this workflow illustrates the strength of our part collection. Its modularity and compatibility allow for flexible engineering of diverse functions, while its adaptability ensures that future iGEM teams can extend the system to target new therapeutics, chassis organisms, or host environments.
These transcriptional units were designed as modular inserts and assembled into the pJUMP backbone using Golden Gate Assembly (GGA). Each construct combines a fluorescent protein (fuGFP or mCherry) or a biosynthetic enzyme (TPL) with specific tags and signals that enable secretion, uptake, or cleavage functionality. To assess the role of each feature, we also created control constructs in which individual tags were omitted. This design allowed us to evaluate the contribution of each element.
Together, this part collection provides a versatile platform for both proof-of-concept (using fluorescent proteins) and therapeutic applications (such as L-DOPA biosynthesis).
In our GGA level 1 assemblies, geneblocks 1–8 were individually cloned into the main site of the pJUMP backbone. In level 2 assemblies, we combined geneblock 8 (mCherry) with each of the other constructs (1–7), enabling us to generate bacteria that simultaneously expressed red fluorescence while secreting GFP or producing L-DOPA (see engineering cycle - cloning - Plasmid design TU).
The constructs were built from well-characterized parts taken from the iGEM Registry, UniProt, or literature. By combining these elements into different transcriptional units, we created a modular collection that allows systematic testing of secretion, uptake, and therapeutic production.
Biosafety is a key requirement in our project. We therefore designed and integrated genetic kill switch systems that allow external control and prevent uncontrolled bacterial growth. The kill switches were cloned into the downstream sites of the pJUMP backbone (see engineering cycle - design kill switches).
All functional geneblocks were assembled into the Joint Universal Modular Plasmid (pJUMP) backbone, a modular vector platform compatible with both Golden Gate and BioBrick standards. Built on the Standard European Vector Architecture (SEVA), the pJUMP collection provides plasmids with diverse replication origins, offering flexibility in copy number and host range, including broad-host-range options .
Figure showing the design of Joint Universal Modular Plasmids from Valenzuela-Ortega & French (2021) .
We made use of a standard cloning strain, our target chassis organism, and a vertebrate model system.
See engineering cycles: Table 1. Parts planned to use from iGEM distribution kit in cycle 1: Assembly using the iGEM distribution kit.