Team:XMU-China/Human Practices
This year, our program focuses on the safety in food production. Considering the local situation and brainstorming in the team, we decided to solve the problem of herbicide residues in tea production. First of all, we interviewed the practitioners in different production links in the tea industry, and they provided valuable opinions for the design and implementation of our project. In the process of project design, we exchanged views with professionals from universities and government departments, covering different fields such as biology, environment and ecology, and mathematics. At the same time, we also kept close contact with other universities and shared a lot of valuable information. In order to integrate our project into the society and promote the ideas and methods of synthetic biology, we conducted public education on synthetic biology through offline activities and distributed questionnaires related to the subject through online channels. In the process of all the above activities, we have ensured the information security of the participants according to the guidance of professionals.
In the whole HP activity, every step complements each other and forms an organic whole.
In our HP program design, every activity has a clear goal. This purpose-driven design method can make the most of HP's results to improve our project. In the process of advancing the project, we divided HP objectives into four sections according to the types of tasks: background investigation, project design, modeling improvement and result feedback, and take them as guidance to carry out activities. Besides, we also attended several communication meetings to listen to more suggestions. Most importantly, in order to ensure that our HP activities can conform to norms and ethics, we consulted relevant people to give guidance on the design of our activities.
Background
Fujian Province, where Xiamen University is located, is one of the most famous tea producing areas in China. We, XMU-China, want to pay more attention to the problems faced by the tea industry when determining the theme this year. In January, 2020, our team went to Wuyi Mountain, Fujian Province for field investigation to understand the current situation of tea pests and agricultural residues.
Team members are communicating with Director Zhong
Team members visited Teacher Da Huo of Wuyi University, Agronomist Shixiong Huang of Wuyi Mountain Agriculture and Rural Bureau and the Director Xingwang Zhong of Wuyi Mountain Tea Quality Inspection Institute. Through the communication with Mr. Huo, we learned that many effective methods such as physics, chemistry and biology have been developed to solve the problems of pests and diseases in tea. By communicating with Mr. Huang, we learned more about the advantages and limitations of biological pesticides. In a word, the vitality of tea trees is extremely tenacious, diseases and insect pests are not the main problems that plague tea fields, but the greater hidden danger lies in herbicide residues.
Aiming at the problems of herbicide application and residue, we are more inspired by Director Zhong. On the one hand, the export of tea and the long-term development of tea gardens need to control the problem of herbicide residues in time; on the other hand, some herbicides such as glyphosate used in tea planting at present do not even have national standard detection methods. If we can develop a convenient detection method for herbicides with incomplete detection standards, it is also a good direction.
This investigation has enabled our team to have a deeper understanding of Wuyi Mountain tea culture and tea industry, and provided great power for us to devote our attention to the development and use of synthetic biological methods to detect glyphosate residues in tea.
In addition to tea farmers, we also visited local tea shops in Wuyi Mountain, and learned about the quality situation and quality inspection information of tea in circulation in the market through the description of the shops. On the evening of August 4th, we visited two tea shops, Shengzhifeng Rock Tea Factory and Beiyuan Rock Tea Factory respectively, and learned that individual tea sellers implement a sampling inspection system for the quality inspection of products sold daily. Once the products sampled by the quality inspection department fail to pass the inspection, they will face the penalty of confiscation of tea products, a certain fine and public notification. At present, merchants basically do not have self-inspection equipment. This also sends us a message: the excessive herbicide residues will do great harm to the interests of the merchants, but the existing methods make it difficult to test by themselves, which makes us more determined to develop a convenient and low-cost testing equipment. As for our subject, they said that they may face certain popularization difficulties at the initial stage of product application, but if there is national authority certification and the safety is guaranteed, consumers will naturally recognize it, which has considerable application prospects.
Visit Shengzhifeng tea factory
On July 22nd, we visited Zhengkui Hu, a local tea farmer in Wuyi Mountain City, Fujian Province, and consulted on the use of herbicides in tea planting and the current weeding methods.
Interview with tea farmer Zhengkui Hu
We were told that glyphosate is a cheap herbicide with high weeding efficiency, which was widely used in tea planting 7-8 years ago. However, in recent two years, with the national attention to herbicide residues and the strengthening of policies related to herbicide use, its use scale began to decrease.
Scene of tea garden after weeding by chemical herbicide
Field visit to tea garden
We were also told that manual weeding and weeding with weeding machines are the main weeding methods at present. Although these physical weeding methods can reduce the damage to the ecology of tea gardens, they also face the disadvantages of poor weeding persistence and high cost. After listening to our topic introduction, Hu was very interested and expressed the opportunity to use our synthetic biology system in his own tea garden. In addition, we also interviewed tea farmers in Qingshiyan Tea Factory by telephone, and they provided similar information.
Design
When the preliminary design of the project was completed and the initial experiment was carried out, the team visited Professor Quanlong Li from the State Key Laboratory of Offshore Marine Environmental Sciences, School of Environment and Ecology, Xiamen University on the morning of July 31st. The professor has been working hard to prepare different testing equipment for marine application. The team introduced to him the problems encountered in the project design and consulting hardware design of the team this year, in order to seek suggestions for improvement.
Team members introduced the project design to the professor
The professor put forward two suggestions on the detection scheme: first, pay attention to the selectivity of reaction in the detection process. Furthermore, we should fully investigate the substances that may interfere with fluorescence signals in tea soup samples. Different kinds of tea water have different types and contents of interfering substances, and then we need to choose the appropriate pretreatment scheme under the background of full investigation.
Our team introduced the design of hardware to the professor
Finally, the professor expressed appreciation and affirmation for the team's projects and led the team to visit various testing projects being researched in his laboratory, teaching the team to consider all details in hardware design before moving from prototype to finished product.
Group photo of our team and professor
On August 3rd, we visited Professor Zhenbin Gong at the State Key Laboratory of Offshore Marine Environmental Sciences, School of Environment and Ecology, Xiamen University. Ask him about our problems in hardware design and fluorescence detection.
Our team introduced the project design to the professor
After communicating with Professor Li Quanlong, we further identified the key issues related to fluorescence detection. The first is the pretreatment of the sample. Professor Gong first gave a general introduction to the detection of agricultural residues in tea. As glyphosate is a target substance with less content, the possible influence of matrix with higher content such as pigment, tea polyphenols and some amino acids in tea should be considered in the detection. It may be difficult to avoid interference by using simple extraction methods. Using molecular stress materials or magnetic nano-materials as fillers, or using the specific combination of some characteristic groups in glyphosate with metal elements are some means worth considering. The improvement of material selectivity will significantly improve the effect of the final product.
In terms of detecting elements, sensitivity and wavelength should be considered. Professor Gong suggested us to use existing commercial instruments to test some low-concentration samples to test whether they can meet our requirements. At present, commonly used silicon photodiodes can basically meet our needs. At the same time, we can consider using ultraviolet photomultiplier tubes or combining multiple laser diodes.
Professor Gong has certain exchanges and cooperation with some government departments, and he enthusiastically expressed his willingness to provide assistance for our further research and continue to communicate with the team.
In order to further promote the degradation of the subject, XMU-China visited Professor Jianqiang Su at the Institute of Urban Environment, Chinese Academy of Sciences. After a brief introduction of our subject, we asked the professor about the use of antibiotics in live bacteria detection, antibiotic labeling and detection.
Our team member is discussing with professors
After learning that the chassis bacteria used in our project are Escherichia coli and the antibiotic used is chloramphenicol, the professor said that Escherichia coli has more natural resistance, and there may be chloromycete-resistant bacteria in the soil, and the penetration of antibiotics into the soil will also bring corresponding potential safety hazards, which are all problems that need to be considered thoughtfully in our project. Therefore, the professor introduced the kill switch module designed to solve the above problems, so as to further ensure the biosafety of our project.
Model Improvement
In the experimental design, the team found that formaldehyde also has a certain inhibitory effect on the growth of cells, but according to the design of degradation kill switch, when glyphosate exists, cells can metabolize to form formaldehyde, and the toxin protein is not expressed, so the cells can survive. Therefore, when formaldehyde concentration is too low, cells will die. When formaldehyde concentration is too high, it will cause damage to cells. So, it is necessary to model the relationship between formaldehyde concentration and cell survival. Through modeling, we can determine the optimum formaldehyde concentration of engineering bacteria (which corresponds to glyphosate concentration at last), and it is also convenient to find suitable culture conditions in the future industrialization. In order to seek advice on modeling degradation kill switch, our team visited Associate Professor Chuanyi Yao, who focused on the simulation and optimization design of protein reaction and separation process in School of Chemistry and Chemical Engineering.
The team introduced the current design idea of modeling to the professor. After understanding the preliminary modeling equations, the professor suggested that the equations should be integrated and optimized, with each protein corresponding to one equation, ignoring the intermediate reaction steps, thus obtaining three groups of relationships between formaldehyde and repressor protein, repressor protein and toxin protein, toxin protein and cell survival. In addition, the effect of formaldehyde on cell growth should also be considered in the equation. The monad equation can be used to find out the effect of formaldehyde on cell growth, determine the effect of toxin protein on cell growth, and determine the parameters of the equation. finally, the correctness of the equation is verified by experiments.
The professor's suggestions greatly enrich and supplement the modeling design of this part, and clarify the key points of modeling work.
This year, our team communicated with CSU-China on the modeling of mutated PhnJ, and found an easy way to compute the binding energy between our mutation and the endogenous C-P lyase.
Share our project with CSU by live-streaming
Since we can’t use the server of our school to calculate the binding energy of the different subunits of the enzyme, CSU-China recommended us using Rosetta to evaluate the binding ability of the endogenous C-P lyase and exogenous PhnJ subunit. This enabled us to model on personal computer equipped with normal hardware.
Online meeting with CSU
Also, CSU-China suggested us rent a server to run MD on it, this could enhance the accuracy of the model.
According to the content of our project, we issued a questionnaire to investigate the social response to our project.
Our questionnaire investigated the public's understanding, attitude towards pesticide residues in tea products, attitude towards our project and some consumers' consumption habits of tea products. We have collected 262 questionnaires, mainly from the consumers of tea products. Now, we have selected some of the survey results of our questionnaire for display. (The question number of each picture corresponds to our original questionnaire.)
1. Consumers’ understanding and attitude of pesticide residues
It can be seen from the above results that most consumers have heard about pesticide residues in tea products. For the types of residues, the most frequently heard by consumers are insecticides and herbicides. Therefore, herbicide residue in tea products is a problem that attracts public attention.
However, as can be seen from the above figure, although most consumers have heard about the problem of pesticide residues in tea, nearly 40% of the respondents still ignore this problem when buying tea. After paying attention to this problem, a small number of consumers will choose to buy tea products of specific brands. The remaining 40% of consumers have a strong attitude, saying that they are well aware of this situation and think that this phenomenon should be solved as soon as possible.
It can be seen from the above figure that consumers have a firm attitude towards pesticide residues. The vast majority of respondents believe that it is necessary to set limits on the use of various pesticides in tea planting. At the same time, strict detection is necessary for herbicide residues with carcinogenic potential such as glyphosate.
2. Consumers’ attitude towards our project
3. Consumption habits of consumers
As can be seen from the above figure, on the premise of ensuring safety, most consumers hold a positive attitude towards our project, and some consumers may hold a positive attitude after considering the cost issue. Assuming that our project can become a trend, most consumers are willing to change their attitudes and avoid the potential risks brought by glyphosate through our project. This reveals two messages to us: firstly, the content of our project is welcome; secondly, only by ensuring the safety of the project can it be widely accepted.
Finally，it is not difficult to see from the above figure that when consumers choose tea products, they first look at the source and seller of tea.
Besides consumers, tea farmers are also our main survey objects, so we designed another questionnaire for tea farmers. This questionnaire mainly investigates two issues of defense: tea farmers' attitude towards herbicides and tea farmers' attitude towards our project. We have collected 262 questionnaires. (The question number of each picture corresponds to our original questionnaire.)
1. tea farmers’ attitude towards herbicides
As can be seen from the above figure, more than half of the tea farmers surveyed have admitted to using herbicides, but most of them said that they have stopped using herbicides now.
An interesting phenomenon can be found from the above picture: only about a quarter of the respondents think that herbicides can improve the yield and quality of tea. But when asked what negative effects will be caused by not using herbicides, many tea farmers think that this will greatly increase the labor cost. It can be seen that herbicides play a great role in cost control in the production process.
However, from the above figure, once the herbicide residue of the product is found to exceed the standard, the tea farmers will face the risk of losing their reputation and market competitiveness. Therefore, the use of herbicides is in a dilemma in tea production.
From the above figure, when we mentioned that there is a certain technology that can quickly detect herbicide residues in tea products, all tea farmers expressed their positive attitude towards this technology. However, when we said that the technology was based on genetically modified engineering bacteria, half of the tea farmers said they would no longer support this technology. This intuitively shows that there are still some misunderstandings and prejudices about synthetic biology.
Fortunately, however, when we explained to the respondents that our method was safe, most tea farmers showed a positive attitude towards our project again. On the one hand, it shows the importance of maintaining accurate information exchange with the society during the project advancement, on the other hand, it shows the importance of safety for our project. At the same time, it also shows that the advantages of synthetic biology in solving some problems can be accepted by people only after the public misunderstanding of synthetic biology is eliminated.
Communicating Meetings
Due to the COVID-19 epidemic, this year's South China Regional Exchange Conference was held in the form of online conference, and more than 30 teams from South China introduced their projects through video on social media. After the meeting, we reached an in-depth exchange intention with Central South University, Xi'an Jiaotong University and Sichuan University, got a detailed understanding of each other's subject content, and put forward many useful suggestions to improve each other's subject design. In the follow-up work, Central South University has further cooperated with us in mathematical modeling and physical modeling, and the information shared by both sides has greatly accelerated the progress of modeling work of our two teams.
CCiC is the abbreviation of Conference of China iGEM Community. This year's seventh CCiC could not be held offline due to the epidemic situation. Sixty teams attended the conference on August 28-29 through online conference and video display, and shared their respective topics. During the meeting, we actively maintained communication with other teams, and gained many valuable suggestions while making suggestions.
The team members are participating in the discussion of the online meeting
After the project exhibition, the judges put forward helpful suggestions on our presentation method, and affirmed our report effect.
Participating in CCiC and getting the best poster award
In order to further understand the official requirements of iGEM for the informed consent of collecting personal information and opinions, and to promote the implementation of Human practices, we interviewed M.D. Bocen Lin from School of Public Affairs of Xiamen University, she mainly participated in the survey project of disabled families in Fujian Province, and visited Sanming and Zhangzhou in Fujian as an investigator.
Team member is discussing with M.D. Lin
In terms of the "informed consent" of the respondents, M.D. Lin summed up two key points that need to be clarified before conducting the investigation: first, clarify the purpose of the investigation and the required data; Second, clarify the identity of respondents, such as government officials, ordinary citizens, or the division between urban and rural residents, so that they can explain the issue of informed consent easily and easily in different ways. Whether it is questionnaire survey or recording, investigators need to ensure the respondents' right to know the purpose and content of the survey and ensure the safety of their collected personal information.
With regard to the certification Committee of the "informed consent form", she said that there are few professional social investigation ethics committees in colleges and universities at present. Generally, the research work of teachers and students in colleges and universities is mainly carried out by issuing a research letter, and the unit agrees to accept the research and help arrange the relevant research work, and then goes to the place to carry out on-the-spot investigation. Both the research letter and the consultation letter are approved by the official seal of the college. It is suggested that we submit the informed consent form as an annex to the research letter to the college for examination and approval, and also issue and explain it to the interviewees during the research process.
In addition, in view of other problems that may be encountered during the investigation, M.D. Lin made supplementary explanations on the contact work in the investigation and the way to ask sensitive questions. Through the communication with M.D. Lin, our doubts about the informed consent of collecting personal information and opinions have been solved. At last, M.D. Lin affirmed our current work and the rigor of our research, we also thanked her for answering our questions. Through training, we have strengthened our belief that excellent HP activities are based on full respect for others and their information.
Our HP is not a loose group of social activities independent of the project. At the beginning of the design of HP scheme, we considered how to combine it with the design and promotion of the project. After the discussion in the team, we determined the basic idea of HP: improve the subject design through the information obtained by HP, guide the experiment through the subject design, and finally evaluate the effect of HP by the test results. At the same time, the public opinion is also very important to us, so after perfecting the project design to a certain extent, we immediately designed a questionnaire to collect feedback from the society.
Of course, all our HP is strictly in accordance with the relevant safety guidelines.
Background
Since the high-rise buildings rise from the ground, we know that the mastery of the basic information of the project will directly affect the completion quality of the project. Therefore, we have communicated with authoritative departments in agriculture and tea industry, traced back to the production and sales of tea products, and directly communicated with tea farmers and retailers to understand their needs. Through communication, we understand the urgency of solving the problem of agricultural residues in tea products and developing simple detection methods. These two pieces of information undoubtedly provide good support for our project-our project is meaningful and necessary, and once realized, it will meet the needs of specific people and fill the relevant gaps.
Based on the information obtained by HP and brainstorming within the team, we determined the specific direction of this project: using synthetic biology to construct a simple method for detecting glyphosate residues in tea products, and developing a synthetic biological method for degrading glyphosate during tea planting.
Design
In order to improve the design of the subject content and provide reliable guidance for the experimental design, we have made several dockings with professionals in universities and scientific research institutions. In every interview, we insist on problem orientation and fully consider how to improve the positive impact of our project on society. Professor Quanlong Li pointed out the problem of fake positive in detection and proposed to use anchoring system to avoid errors. Professor Zhenbin Gong assisted us in the selection of hardware components, and put forward constructive suggestions on the construction and popularization of hardware. Professor Jianqiang Su put forward some suggestions on biosafety in our subject, and suggested that we introduce kill switch system.
The HP activities in this link are advancing synchronously with our experiment, and the combination of experiment and HP is achieved. After discovering the problem, help us solve it through HP, which is in good agreement with our general thinking of HP this year. Therefore, the information obtained in this stage is directly reflected in our project design: in the detection direction, the protein used for detection is anchored on the surface of engineering bacteria; In the direction of degradation, the hardware was constructed according to Professor Gong's instruction, and kill switch was used in engineering bacteria according to Professor Su's suggestion.
Model Improvement
At the beginning of the project design, we have a preliminary design for modeling. With the advancement of the experiment and the further improvement of the project design, we determined two modeling directions: protein molecule docking and kill switch related modeling, and carried out HP activities in these two aspects. After understanding our modeling objectives, Professor Chuanyi Yao helped us to deduce the modeling equation, and helped us to clarify the relationship between toxic protein, formaldehyde and cell growth during kill switch operation. In addition, through cooperation with Central South University, we fully exchanged views on mathematical modeling and molecular docking, which promoted each other's work progress.
With the help of Professor Yao, we perfected the idea of mathematical modeling, and finally deduced the corresponding equations. Through the equation, we can get the threshold value of formaldehyde concentration that causes bacterial death, thus determining the working effect of kill switch. The results of this work also inspire us to improve the formaldehyde promoter and enhance its sensitivity to formaldehyde response, so as to enhance the sensitivity of kill switch. At the same time, by analyzing the results of molecular docking modeling, we designed and mutated the key enzyme phnJ to improve its ability to degrade glyphosate.
Results of Our Questionnaire
Through the analysis of the questionnaire results, we know the social understanding and attitude towards pesticide residues in tea products. These analysis data are consistent with our initial interview results for tea farmers and tea merchants, which proves that our research is meaningful and necessary.
The analysis results of these questionnaire data do not directly affect our project design, so we have no way to give direct feedback to it like to the interview results of experts in our design. However, we used these valuable social survey data when establishing cooperative relations with enterprises.
In order to make full use of the information from the questionnaire, we used the information to communicate with tea companies. Based on the analysis of the information, we both reached a consensus and finally reached a cooperative relationship. Please refer to the "Business Cooperation" section for more details.
Value orientation of the project
In the process of project promotion and improvement, we always put safety in the first place. The vision of our project is to promote the use of synthetic biological detection and degradation of glyphosate on a large scale in the society. Once these new methods are used on a large scale, any slight potential safety hazard may turn into a problem that seriously endangers public safety. Therefore, the demand for safety is at the highest position in our project. This is reflected in our HP and modeling work in kill switch, and this problem is clearly explained in the questionnaire distributed to the public.
When the project is implemented, we hope to protect the benefits of users (tea farmers, tea merchants, etc.) to the greatest extent. This point can be fully reflected by our HP design: we have adopted many suggestions from tea industry practitioners. And in the process of HP, we were also moved by the industrious and honest quality of these workers, thus strengthening our previous ideas.
Because our project uses mature technology and does not directly edit the genes of animals and plants, it will not introduce ethical issues.
Close the loop!When carrying out HP activities, we pay great attention to translating other people's suggestions into the design in our subject, thus forming a closed loop between other people's expectations and our design. These positive feedbacks are shown in the following figure.
Business cooperation
In order to apply the results of the project "An-TEA Glyphosate" to the actual production of tea in the future, we decided to set up a company whose main business is to produce products that can detect and degrade glyphosate in tea. We have written a business plan (If visitors of this webpage want to browse details of the questionnaire, please click the link supporting Entrepreneurship) and hope to find a partner.
On October 2, we visited Qingshiyan Tea Co., Ltd. and Wuyishan Ruizhuang Tea Co., Ltd. Among them, Qingshiyan Tea Co., Ltd. was founded in 1926 and was recognized as a well-known trademark in northern Fujian in 2012. We showed them the results of our project and the team's business plan. In the end, the two sides reached a follow-up intention of deep cooperation: Shiyan Tea Co., Ltd. and Wuyishan Ruizhuang Tea Co., Ltd. will provide experimental tea gardens to the team, and in return, the XMU-China team will provide the other company with the first batch of products that free of charge and guide their use.
Xiamen University, Fujian, China
No.422, Siming South Road, Fujian, P.R.China 361005
