Collaborations

Our team is designing an educational experiment that uses fluorescent proteins for painting as a way to stimulate young people's interest in the life sciences.

However, since we are a brand new team and have no reserve in this area, we chose to collaborate with multiple teams and have other teams provide us with the corresponding plasmids, and we can help them characterize the plasmid originals for validation while completing our painting experiment.

We have been helped by BNDS-China, iBowu-China, and BIT-China in this process, and we are also ready to share our acquired plasmids and characterization results to SHSBNU-China.

GFP

Because we are in our first year of participation, we do not have the parts package that teams received in previous years, so we do not even have the basic GFP plasmid. This was the first problem we encountered.

Finally we looked for the empty vector plasmid of GFP where in BNDS-China, sfGFP-pSB1C3,Thanks to their generosity.

We performed transformation and expression after receiving the plasmids. The strain used was BL21 receptor E. coli, and after coating the plate and overnight culture, we successfully obtained the strain expressing green fluorescent protein. We sent back this result and informed them about our expression conditions.

RFP

Regarding the source of RFP，the iBowu-China team gave us a clue and their mentor told us that we have the corresponding plasmids from BGI and also helped us to contact the corresponding person in charge. Thanks to their generosity, they gave us mCherryFP directly instead of RFP. Here the process of validation and transformation was saved for us, and we just proceeded to shake the bacteria directly.

BFP

In the process of searching for the final BFP, we found that SHSBNU-China was also searching for the BFP, and we learned from the clues they gave that BIT-China had used the BFP in previous years, so we met the captain of BIT-China through the WeChat group in China, and the captain of BIT-China kindly sent us a plasmid It is a plasmid that contains the mTagBFP plasmid.

Looking at the DNA profile of this plasmid, we found that there was no promoter on this plasmid, so we designed our own primers to obtain the mTagBFP part of the gene and then transferred it to the pet28a plasmid that we used in this year's experiment. fluorescence part can be expressed in BL21 with the induction material IPTG.

After completing the plasmid reconstitution experiment, we transformed BL21 with the new plasmid, T7-BFP-pet28, and found that the colonies grew very little after coating the plates. After extended incubation, the following LB plate was obtained. After that, a single colony was selected and induced with IPTG after incubation in LB medium and excitation at 399 nm and measurement of emission light at 454 nm showed that the result did not reveal fluorescence. We think it may be that the plasmid reconstitution failed.

So unfortunately, we didn't get a usable BFP in the end, and we are going to continue to optimize the original plasmid and send it for sequencing at a later stage to determine whether our construction is successful. We also gave feedback to BIT-China about this uncharacterizable result, and we learned in our communication that they did not succeed in expressing this plasmid even after acquiring it, so the promoter was not added to the plasmid.

Summary

For the final educational activity, we are using both GPF and mCherryFP plasmids for transformation and plate coating. Thanks again to the three teams BNDS-China, iBowu-China, and BIT-China. If other teams would like to try the fluorescent bacterial painting experiment, we can provide suitable plasmids and bacterial fluids for you.