How successful was the team in engineering a plant or algal cell?
The final results of engineering for our plants are yet to be determined. A floral dip transformation on our Wisconsin FastPlants (Brassica rapa) was carried out on 10/10, and supposing that the seed pods mature in time for the Jamboree, we hope to present more results during our judging session. To validate our engineering success, we’ve planned four experiments:
Does their work address a need or problem in plant synthetic biology?
Our work addresses the problem of growing plants in space, where plants are unable to direct root growth downward because there is no gravity to force statoliths to direct gravitropism. We aim to replicate this mechanism by localizing ferritin to the statoliths of A. thaliana and using a magnetic field gradient to direct the root growth downwards. This research could contribute to advancements in making plant growth more efficient and investigating factors that optimize plant growth.
How well did the team use the special attributes of the plant chassis?
We used Arabidopsis thaliana and Brassica rapa as our plant chassis and by using the statoliths involved in the gravity sensing in the root and instead generating a magnetic field so that the increased ferritin molecules would instead be pulled down. Other plants grow far slower than A. thaliana and this specific chassis allows us to study the movement of roots in media and as it grows.
Are the parts/tools/protocols for plants made during this project useful to other teams?
The 2022 Hopkins iGEM team wants to make it easier for teams to perform experiments with applications in space and microgravity. In order to effectively simulate microgravity on earth, a clinostat, or random positioning machine, is essential. Team Concordia-Montreal developed an excellent 3D clinostat in 2021, which rotates in 2 axes to average the gravity vector in X, Y, and Z. Some experiments that are constrained to a flat surface like an agar plate may not require the additional electrical and mechanical complexity of a 3D clinostat. To this end, we created an open-source design for a 2D clinostat to achieve random positioning of agar plates, and is usable for microgravity experiments including plants. Our design has nearly identical functionality, but most parts are 3D-printable, the stepper motor is more inexpensive, and design files are accessible on GitHub.
Plants at APL
We collaborated with Jeff Shipp from JHU’s Applied Physics Laboratory (APL), who graciously grew our plants for us and assisted us in Agrobacterium transformation and floral dip transformation. Arabidopsis had been planted earlier this Fall, but unfortunately due to delays in our plasmid assembly and verification, the window for transforming our plants had passed. As a result, we transformed a batch of Wisconsin FastPlants, a fast-cycling variety of Brassica rapa developed at UW Madison. More specifically, we used the AstroPlants strain, which has a dwarfed phenotype and has been used in on-board NASA experiments. Initially, we grew them for two weeks in a greenhouse space, but temperature, light cycle conditions, and nutrition were not optimal, and the plants only grew to an effective maturity of 5 days. Once provided with adequate nutrition and a 24-hour light cycle at APL, these plants grew to flowering after an additional 12 days.
Agrobacterium transformation of our five successful plasmids (for the transit peptides prcpHsc70-1, prSSG1, prSS4, prSGR9, and prDPE1) was successfully performed using electroporation at APL on 9/28.
Floral dip transformation, a transformation method in which the above-dirt portions of the plant are dipped in Agrobacterium liquid culture, was performed on two plants for each of the five plasmids at APL on 10/10.
