Overview
Being high school students, human practice activities have played an important role in the successful completion of our project. Human practices broaden our knowledge, expand our thought, and clarify the purpose and content of our project. All presented on this page is how we improved our iGEM project through human practice activities, and confirmed that we are going to develop a detector used to monitor pathogenic bacteria in refrigerators, reminding people to clean and disinfect refrigerators in time, preventing diseases caused by fridge bacteria, and keeping peoples physically healthy. In addition, human practice activities have played a huge role in smoothing progress of our iGEM project.
1. Online survey questionnaire
After several times of brainstorming activities, we are going to develop a detector to monitor pathogenic bacteria in refrigerator in our project. In order to broadly understand people’s concern about this issue, we designed a survey questionnaire and put it online to obtain responses from people who could use smart phone. Some mobile screenshots of the survey work online (the last update is on July 30th 2024) are shown as follows (Fig.1).
Fig.1 General information of online survey questionnaire
We designed some relevant questions in the questionnaire, and some important questions’ responses are shown in the followings:
The first question is that “Do you know anything about bacteria in refrigerator before?” From the result we can see that 57.32% of the respondents know about it clearly to some extent, and 32.56% of people know a little bit about it (Fig.2), indicating that most people are very clear to bacteria in refrigerator.
Fig.2 Responses to the question “Do you know anything about bacteria in refrigerator before?”
(a) Yes, it’s clear to some extent; (b) Know a little bit; (c) Have no idea.
The second question is that “What do you think are the harms of bacteria in refrigerator? (multiple choices question)" All the choices are right answers, but from the results we can see that most people cannot select all the right choices (Fig.3), suggesting that the respondents know some harms, but they do not understand completely the symptoms caused by bacteria in refrigerator.
Fig.3 Responses to the question “What do you think are the harms of bacteria in the refrigerator? (multiple choices question)”
(a) Diarrhea; (b) Vomiting; (c) Fever; (d) Headache; (e) Abdominal cramps; (f) Nausea.
The third question is that “What diseases do you know could be caused by bacteria in refrigerator?” This is also multiple questions since bacteria in refrigerator could cause many diseases. All of the choices are right, but the response of this question shows that no one selected all the disease, and most people only know 1 or 2 diseases (Fig.4).
Fig.4 Responses to the question “What diseases do you know could be caused by bacteria in refrigerator? (multiple choices question)”
(a) Bacterial dysentery; (b) gastroenteritis; (c) meningitis; (d) appendicitis; (e) sepsis; (f) typhoid fever.
The next question is that “How often do you clean your refrigerator?" The result showed that 42.55% of respondents clean the refrigerator every six months and 31.92% (c and d) of respondents clean the refrigerator ONCE more than a year (Fig.5), indicating that many people overlook the cleanliness of their refrigerators.
Fig.5 Responses to the question “How often do you clean your refrigerator?”
(a) Once a month; (b) Once every six months; (c) Once a year; (d) Once every two years or more.
Summary for survey work online: (1) Many people know that there may be some bacteria in refrigerator; (2) Most people know some of the harms and diseases caused by bacteria in refrigerator, and they don’t understand this problem very well; (3) Many people do not often clean the refrigerator.
These results of survey work online strengthened our determination to do something for solving the bacterial contamination problem in refrigerator in this iGEM competition.
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2. Interview with Dr. Fan of gastroenterology of Qianfoshan hospital
To understand comprehesively the impact of pathogenic bacteria in refrigerators on people’s health, we interviewed with Dr. Fan of gastroenterology in Qianfoshan hospital. Dr. Fan is an expert in gastroenterology who is very familiar with diseases caused by gastrointestinal pathogenic bacteria. He was so busy that the interview activity was conducted in the doctor's duty room.
Fig.6 Interview with Dr. Fan of gastroenterology in Qianfoshan hospital.
He was so busy that the interview activity was conducted in the doctor's duty room.
Here are summarized some contents of the interview with Dr. Fan:
(1) Situation:
In fact, there are many bacteria in refrigerator (Fig.7). The types of pathogenic bacteria in refrigerators mainly include Shigella, Salmonella, Escherichia coli, Listeria, Yersinia, and so on.
Fig.7 A large number of pathogenic bacteria in refrigerator.
Dr. Fan also introduced that the cases caused by fridge bacteria accounts for 20%-30% in all clinic patients every year, usually occur in spring or summer. The symptoms of refrigerator bacterial poisoning are diverse, but usually mainly gastrointestinal symptoms such as nausea, vomiting, abdominal pain, diarrhea, etc. In severe cases, symptoms such as dehydration and fever may occur. For specific bacterial infections such as Listeria monocytogenes, symptoms similar to upper respiratory tract infections (such as chills, fever, headache, muscle pain) and central nervous system infections (such as meningeal irritation, ataxia, etc.) may also occur.
(2) Prevention:
Currently, there is no practical method to prevent diseases from the pathogenic bacteria in refrigerators. The only method is to keep the fridge clean, regularly disinfect it. These are crucial for reducing pathogenic bacteria in the refrigerator.
However, the reality is that people often start cleaning their refrigerators only when they smell unpleasant odors inside. Actually, at this point, the bacteria in the refrigerator have already begun to multiply in large numbers, posing a serious threat to people’s health, even life.
Opportunity and challenge: From the interview with Dr. Fan, we figure out that it is important that people need a something device in refrigerator to detect bacteria, telling people when is the time to clean or disinfect the refrigerator for preventing some diseases. If we develop a method to detect bacteria in refrigerator, some injuries would be avoided.
It is an opportunity and challenge, making us to make a decision that developing a testing method for detecting bacteria in the refrigerator in our iGEM project.
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3. Interview with professor Liu of Shandong University
After making our decision, we need to know more knowledge about how to detect bacteria in the refrigerator. What are the characteristics of pathogenic bacteria in the refrigerator? What are their targets for detection? What are the methods of detection? And what are the precautions, etc. We were confused and disoriented with many questions although we browsed a lot of literature. To broaden our knowledge, bringing some questions, we interviewed with professor Liu Zhifang who works in Shandong University. Due to the working time arrangements, we held a virtual meeting with Dr. Liu (Fig.8).
Fig.8 Interview with professor Liu in virtual meeting.
First, Prof. Liu highly praised our project. She said that it has special significance to develop a test method of bacteria in refrigerator, reminding people to clean and disinfect refrigerator.
According to our demand, professor Liu introduced some bacteria detecting methods. She told us there are 3 common bacterial detection methods based on molecular recognition: recognition of nucleic acid probe, antigen antibody immune, and nucleic acid aptamer ligand, which are showed in Fig.9. And she introduced the mechanism of these 3 bacteria detection methods.
Fig.9 Bacterial detection methods based on molecular recognition.
At last, Prof. Liu suggested that combining aptamer with reporter gene to construct a toehold swithc is a feasible method for detecting bacteria (Fig.10).
Fig.10 Priciple of bacteria detected by the RNA Toehold Switch.
After interview with Prof. Liu, we searched and found the aptamer sequences of Shigella dysenteriae, Escherichia coli, and Salmonella typhimurium. According to these sequences, we designed the bacteria detecting method, using amilCP, mRFP, and EGFP as reporter genes to display the testing result on the biosensor.
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4. Face to face survey in community
After finishing the designation of our project, we started the wet experiment in the lab. However, we also worried about if people really need such a device to detect bacteria in fridge? Did people would like accept our detector? In order to address these doubts, we set a paper questionnaire and performed some survey work face to face in community to explore significance of our project (Fig.11).
Fig. 11 Interview with people face to face in community.
From the face-to-face survey questionnaire, the responses to some questions are shown as follows:
Responses to the question "Have you ever had a question about whether it's time to clean your refrigerator?" showed that 74.47% respondents said “Yes” (Fig.12), indicating that the majority of people don’t know when they should clean their refrigerators.
Fig.12 Responses to the question “Have you ever had a question about whether it's time to clean the refrigerator?”
(a) Yes; (b) No.
The next question is that "Do you need a device in your fridge to detect bacteria and tell you it’s time to clean the fridge?" 95.74% of respondents select “Yes”, indicating that the bacteria detector in fridge will be popular if it is produced in reality (Fig.13).
Fig.13 Responses to the question “Do you need a device in your fridge to detect bacteria and tell you it’s time to clean the fridge?”
(a) Yes; (b) No.
The third question is that “Do you accept a device to detect bacteria in fridge using synthetic biology?” 51.46% of people response “Yes”, although they don’t know synthetic biology very well, 28.7% of people responded “Maybe accept it”, and 19.78% of people responded “No idea” (Fig.14), illustrating that most people would like to use our product.
Fig.14 Responses to the question “Do you accept a device to detect bacteria in fridge using synthetic biology?”
(a) Yes, although I don’t know it very well; (b) Maybe accept it; (c) No idea.
The last question is “How much price increase can you accept a refrigerator with a bacterial detector?” in the face-to-face survey. The result showed that most people can accept 10% price increase, and the others can accept less than 30% price increase (Fig.15), indicating that our detector would be accepted by all the consumers if its cost is less than 10% price of a refrigerator.
Fig.15 The survey result of question “How much price increase can you accept a refrigerator with a bacterial detector?”.
(a) less than 10% price increase; (b) less than 20% price increase; (c) less than 30% price increase; (d) less than 40% price increase.
Conclusion: From the face-to-face survey in community, we know that:
(1) The majority of respondents didn’t know when they should clean their fridges, and they need a device to detect bacteria in fridge, reminding them to clean or disinfect the fridge.
(2) Most people can accept a detector developed using synthetic biology, and they would like to use the refrigerator with a bacterial detector if its price is less than 10% increase, compared to the normal price of a refrigerator.
These results encouraged us to develop a bacterial detector for use in refrigerators, using synthetic biology methods. But it is important to keep the cost of the detector within 10% of the refrigerator price.
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5. Consulting engineer of Qingdao Haier Biomedical Co., Ltd
Since our product would be used in refrigerators, and the biosensor containing cell-free system hydrogel on which bacteria are detected, changing the hydrogel color to blue, how to design it to be a sensitive and convenient device is important. Bringing this question, we visited Qingdao Haier Biomedical Co., Ltd to consult the design of our detector (Fig.16).
Fig.16 Visiting Qingdao Haier Biomedical Co., Ltd for detector design.
In Haier Biomedical Co., Ltd., they first led us to visit their company's products, one of which is refrigerator, and introduced the design and productive process. At the same time, we introduced our project, telling them we want to design a biosensor to detect bacteria in refrigerator. In the communication with them, the engineer suggested that it should consider the environmental temperature and medium of the biosensor reaction, the location of the settings, and the intuitiveness and visibility of the detection results.
In addition, they suggested that it should be better if we convert the color values on the biosensor into electrical signals, and then amplify and quantify the electrical signals to enhance the accuracy of the detection results. These suggestions are in line with our wishes to a more accurate and more sensitive biosensor (Fig.17).
Fig.17 Passionate discussion with engineers on the design of biosensors.
Refrigerator is one of their main products, and refrigerator bacterial detector can optimize their product performance, so they were very interested in our project. And they expressed the willing to cooperate with us in the future.
From visiting company, we got some inspiration to make a biosensor for detecting bacteria in refrigerator, which is convenient and feasible. They are very interested in our project and look forward to further cooperation in the future.
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6. Consulting Prof. Liu again to solve problems
During the experiment, we encountered many problems. For example, the detecting sensitivity was relatively low using the chromoproteins as reporters. And what kind of medium can be used for bacteria detection in refrigerator?
Prof. Liu suggested that it would be better if we use lacZ marker gene instead of the chromoproteins to display the testing result, because lacZ gene encodes β-galactosidase which catalyzes the enzymatic reaction to amplify the signals. Many different substrates are available for this enzyme. We can use the substrate X-Gal to test β-galactosidase activity, indicating the result of bacteria detection. This is really helpful!
Prof. Liu also introduced hydrogel which is a kind of three-dimensional reticulated gel with high hydrophilicity. Natural polymer hydrogels such as gelatin and chitosan have good biocompatibility and degradability. Hydrogel has become an important matrix material in cell free system due to its unique 3D network structure and good biocompatibility. The application of hydrogel in cell free system is a cutting-edge and rapidly developing field.
She suggested us to try constructing a cell free system hydrogel to detect the pathogenic bacteria in refrigerator.
Under the guidance of Prof. Liu, we determined to use lacZ coding β-galactosidase as the reporter which catalyzes the substrate X-gal to produce blue product, in which the signals are amplified to increase the detection sensitivity. And we tried to construct the cell free system hydrogel as the detection medium. We really appreciated Prof. Liu who gave us a great enlightenment and less detours.
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7. Publicity and education
To better promote the development of synthetic biology and publicize our project, we established a synthetic biology club in International Center of Jinan Foreign Language School (ICJFLS). In the recruitment activities, we introduced the history, development, application and prospect of synthetic biology, as well as iGEM competition. In addition, we introduced the purpose, principle, design and significance of our project (Fig.18). Many new students are interested in these introductions, showing great enthusiasm in it. We hope the new students of our club will participate in iGEM competition next year.
Fig.18 Pictures taken in the recruitment activity of synthetic biology club.
From the survey questionnaire results, we can see that although some people have some understanding of the situation of pathogenic bacteria in refrigerator, they still do not attach enough importance to this issue. And there are still some people who are not aware of the harm or diseases caused by refrigerator bacteria. People also don't know when to clean their refrigerators and how to do so. Therefore, it is necessary to publicize some relevant knowledge to the public.
In addition, from the results of the social survey, we can also see that our project and product have significant social meaning, and the public has a strong demand for our product. In order to let more people know about our product and help it go to the market as soon as possible, we make full use of the modern self-media platform Bilibili and Tiktok account to promote our project and product, making it better serve the society (Fig.19).
Fig.19 We used some self-media platform (Bilibili and Tiktok) to publicize our project.
(A)Home page screenshot of our Bilibili account. (B) Home page screenshot of our Tiktok account.
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8. Conclusion
(1) From the survey work online and face to face, we know that bacterial contamination in refrigerator is a big problem in the public. We decided to do something to help solving this problem using synthetic biology in 2024 iGEM competition. And we know it has a great market demand!
(2) From interview with Dr. Fan of Qianfoshan hospital, we found an opportunity and challenge, making us to make a decision that developing a testing method for bacterial detection in refrigerator, reminding people to clean the fridge in time.
(3) After interview with professor Liu, we determined to use RNA aptamer switch and reporter proteins amilCP, mRFP, and EGFP to detect bacteria in refrigerator.
(4) From visiting company, we got some inspiration to make a hardware and come to understand the importance of detecting signal amplification. They are very interested in our project and look forward to further cooperation in the future.
(5) Visiting Prof. Liu again, we determined to use lacZ coding β-galactosidase as the reporter gene, resulting the signals amplified. Using the cell free system hydrogel is very good choice.
(6) Publicity and education activities made our project more meaningful. We helped the public to know some common sense about bacteria in refrigerator, and they gave us more suggestion and improve our project. We publicized synthetic biology, iGEM and our project to the society.
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