Find out more on how this project started over here. We'll share with you the inspiration behind our brilliant idea and how it grew. You'll also be able to learn the reasons why the team dedicated itself to this project.
The Problem
Earlier in 2019, a dead whale washed up on the shore of the Philippines with 88 pounds of plastic found in its stomach. And unfortunately, according to the United Nations Educational, Scientific and Cultural Organization, this whale is amongst the over 100,000 other marine animals found dead with plastic-filled stomachs.
By the end of 2015, the world had accumulated over 7.8 billion tons of plastic. The most common type of plastic that is polluted into the ocean is found in the everyday single-use plastic water bottle: PET (polyethylene terephthalate). These plastics are eventually broken down into 5 millimeters to 100 nanometer particles called microplastic, through the process of degradation called hydrolysis. And unfortunately, this is a very slow process, as seawater is not a considerable environment for hydrolysis. Because of this, it can take a single PET water bottle to take almost 450 to degrade.
This microplastic pollution poses a dangerous threat to oceanic marine life. One 2013 Nature study found that PET particles in the ocean carried chemical pollutants from the marine environment. Once these plastic particles were ingested by fish, they accumulated these chemicals and went into hepatic stress from suffering liver toxicity. And these microplastics have found their way up the food chain. One National Geographicarticle reported that microplastics were recently found in human feces. Although the full consequences of microplastics on human health specifically haven’t quite yet been fully discovered. However, microplastics have proven to cause nflammation in the lungs once inhaled through from atmosphere. Unfortunately, the problem of plastic pollution will only get worse, as it is currently predictedthat within the next decade, the amount of plastic in the ocean will triple.
Current Solutions
As of now, there are several solutions that were created to combat the problem of plastic pollution, specifically the presence of microplastics. According to The Handbook of Environmental Chemistry, for most scientists and policymakers, thorough oceanic microplastic cleanup is neither logistically nor economically feasible. That’s why one proposed a solution was to introduce more policies with upstream interventions, which involve regulation and economic incentives.
Moving away from policy, scientists have researched more innovative techniques to combat the PET microplastic problem. One project is researching how jellyfish slime could help. The EU-funded project called GoJelly has been examining the mucus of jellyfish and analyzing thier absorption properties. Essential, they are looking into the potential of absorbing microplastics with the mucus. However, more research has to be performed before testing the method. Another potential solution involves the use of nanotechnology. In July of 2019, scientists designed nano-sized reactors that triggered microplastic to breakdown. These reactors are called nanocoils and are also reuseable. However, like the GoJelly project, more research needs to be performed before using the nanocoils in nature. For example, the reactors would need to be testing in a variety of different organic materials, similar to that of seawater
Our Solution
Our solution is something innovative because we are using cyanobacteria in the environment to conduct this process. New research has discovered the bacteria Ideonella sakaiensis which has the capability to break down PET by producing the protein PETase.Our solution involves using the PETase gene and modified it in order to be expressed and secreted in cyanobacteria at its maximum potential. Once secreted by cyanobacteria, PET plastic degradation will occur. Our goal is to clean our oceans, we will be working in the near future for our solution to be approved and released in the marine environment.
Project Description
There are many parts that composed our project. We first have to develop our gene of interest, this work was done by our fall team. Our gene of interest is SuperPetase which was found from a research. We had to make some changes in this gene in order to function at its optimal power and be expressed in cyanobacteria. This genetic information was then imported into VectorBuilder, in which our vector was created.
For more information regarding the development of our SuperPETase gene, please visit our project tab.
We created a new part in the Parts Registry. This part consists of our modified superPETase gene, a TorA sequence and a myc tag. If you’d like to learn more about our part, please visit our part registry.
While research was being done regarding our part, we came across an igem team from 2013 who has also worked with TorA sequence before and we found that very interesting. If you would like to look for their part, please click here.
When then conducted PCR of our superPETase gene followed by the PCR of our pAM fragments which is the backbone of the vector that would be use to transform cyanobacteria. Our pAM plasmids were obtained from UTEX repository.gel electrophoresis was conducted in order to separate our fragments.
We then purified our fragments and gibson assembly and transformation had to be conducted in order to first transform our gene into competent E.coli cells. Below you will see a successful transformation in our E.coli competent cells.
The next step is to purified this colonies, get our gene and transform into cyanobacteria.
In order to test the cyanobacteria for extracellular production of PETase, a unique method of assaying was developed. This entails determination of the size of the nanoparticles of PET. These particles were labeled with fluorescein. The breakdown of PET was quantified through fluorimetry.
Fluorescent microscope
Fluorescence of label particles was observed by both microscopy and spectroscopy. Particles were clearly fluorescent when viewed under a fluorescence microscope, left, figure 6. On the right, figure 7, the fluorescence spectra of fluorescein embedded in PET nanoparticles (blue upper) is comparable in the emission side to free aqueous fluorescein (green, lower), with maximum near 520 nm, but the excitation spectra is significantly blue shifted, with a maximum near 440 nm, compared to a typical 494 nm excitation maximum for aqueous fluorescein. This shift can potentially be used to monitor release of fluorescein as the particles are degraded.
If you would like to know more about how all of our experiments were conducted and the purpose of our experiments, we encourage you to visit our project tab in which you will find an explanation of our experiments and the design of our gene of interest, pAM plasmids, and our nanoparticles of PET.
Figure 3. Images of particles
Citations
Borunda, Alejandra. “This Young Whale Died with 88 Pounds of Plastic in Its Stomach.” National Geographic, 22 Mar. 2019, https://www.nationalgeographic.com/environment/2019/03/whale-dies-88-pounds-plastic-philippines/.
Hornigold, Thomas, et al. “How Cyanobacteria Could Help Save the Planet.” Singularity Hub, 31 Jan. 2019, https://singularityhub.com/2018/06/04/how-cyanobacteria-could-help-save-the-planet/.
Long, Kat. “New Species of Bacteria Eats Plastic.” The Wall Street Journal, Dow Jones & Company, 10 Mar. 2016, https://www.wsj.com/articles/new-species-of-bacteria-eats-plastic-1457636401.
Spence, Edward, et al. “Membrane-Specific Targeting of Green Fluorescent Protein by the Tat Pathway in the Cyanobacterium Synechocystis PCC6803.” Molecular Microbiology, vol. 48, no. 6, Dec. 2003, pp. 1481–1489., doi:10.1046/j.1365-2958.2003.03519.x.
Whitaker, Hannah. “How the Plastic Bottle Went from Miracle Container to Hated Garbage.” National Geographic, 24 Aug. 2019, https://www.nationalgeographic.com/environment/2019/08/plastic-bottles/.
Wüstneck, Bernd. “In a First, Microplastics Found in Human Poop.” National Geographic, 23 Oct. 2018, https://www.nationalgeographic.com/environment/2018/10/news-plastics-microplastics-human-feces/.
Yoshida, Shosuke, et al. “A Bacterium That Degrades and Assimilates Poly(Ethylene Terephthalate).” Science, vol. 351, no. 6278, Oct. 2016, pp. 1196–1199., doi:10.1126/science.aad6359.
