Team:XHD-ShanDong-China/Human Practices
Human Practices
In order to set a clear direction for the future development of our project，we launched the human practice research carefully. On top of experiments, we also need interaction with the world to revise our project and to popularize the ideas of synthetic biology and our project in the society. For these purposes, we have conducted human practices, including questionnaire survey, the interview with experts, and the propaganda.
During a social practice visit to the fermentation plant, one of our team members learned that during the fermentation process, a large amount of cooling water was needed to ensure that the fermentation tank would not overheat. Because the most suitable growth temperature of common microorganisms for fermentation is about 37 ℃. If the temperature is higher than 37 ℃, the growth of bacteria will slow down with the increase of temperature, resulting in industrial production losses. Therefore, the fermentation plant should constantly use cooling water to cool the fermentation tank, which will consume electricity and water. So we are thinking that if we can improve the heat resistance of microorganisms, maybe we can reduce the power consumption in the fermentation industry in order to control the temperature, and then reduce the production cost in the fermentation industry.
With this initial idea, we first contacted Professor He of Huazhong Agricultural University.
He is engaged in the breeding of agricultural microorganisms, metabolic physiology, fermentation analysis and other related fields of scientific research. We had a telephone interview with him. After learning about our preliminary ideas, professor he was very supportive and positive.
First of all, he introduced the structure of the fermentation tank. The large fermentation tank is cylindrical, with bacteria and stirrer inside. There is a jacket layer in the middle of the tank to cool down the temperature by circulating tap water. Through the refrigerator of refrigeration equipment, the tap water is cooled to the jacket to take away heat. If the heat is not taken away, the temperature in the fermentation tank is getting higher and higher, and the bacteria will die.
Then he told us that Escherichia coli is widely used as a production bacterium. If the heat resistance of E. coli can be increased by 2-3 ℃, the production cost will be reduced by about 10% - 20%. If the thermal adaptability of E. coli is properly improved, the temperature requirement of cooling water will be reduced, thus reducing the energy consumption cost of refrigeration equipment compressor.
The expert advice and affirmation given by professor pushed us to carry out the project according to our original intention. Then we did a questionnaire to find out the attitude of the general public towards our idea and their understanding of synthetic biology and the iGEM competition. Some of the questions in the questionnaire showed below. Most people have a certain understanding of synthetic biology and iGEM. Our project can deepen the understanding of the masses, which is of great educational significance.
Part of the questionnaire survey showed that nearly half of the public (49.53%) did not know much about the application and function of bacterial thermal adaptability, which made us need to do more science popularization and education on the application of improving bacterial thermal adaptability when carrying out public science popularization.
For the improvement of fermentation industry production and yield, the general public have the idea to start from the direction of improving strains. (Improved strains by mutagenesis, change the stain by changing the cultivation conditions and change the fermentation environment make up the majority of 46.73%, 55.14% and 56.54% of the respondents） This has a positive effect on the direction of our project.
Question 5 of questionnaire survey
Question 6 of questionnaire survey
Question 9 of questionnaire survey
The instructor told us that in order to improve heat resistance, we need to screen the relevant heat resistance genes. Under the guidance of our tutor, we found the dgep gene through literature search and reading. The DegP protein expressed by DegP is an essential gene for the high temperature growth of E. coli, also known as HtrA (high temperature requirement a), which is responsible for the digestion of abnormal folded proteins in the periplasmic space in high temperature environment, which is very important for the thermal adaptability of E.coli. At first, we hope to use molecular cloning method: to transfer the plasmid carrying the gene of dgep into E. coli to increase the expression of dgep. But in the process of experiment, we found that this method cannot stably improve the gene expression, because E. coli will lose the plasmid after several generations, and it is unable to inherit the heat resistance property by temperature. Then we brainstormed and thought about transferring strong promoters into E. coli chromosomes to up regulate the expression of dgep gene. This method is very common, but it was quickly rejected by the instructor, because it will make dgep become a stable expression of gene, no longer subject to environmental temperature regulation, not intelligent. But in the process of this discussion, it makes our project direction more and more clear. To determine the direction of the project is to obtain a stable genetic heat-resistant strain, while maintaining the adaptation to high temperature.
In order to achieve the goal of our project better, we must use appropriate experimental techniques. Therefore, we contacted Professor Ma, an expert in synthetic biology and systems biology, who provided us with a new idea and scheme to achieve genetic stability by gene editing. He asked us to try to find the regulatory genes that are closely related to the dgep gene. Then we try to up or down regulate the expression of dgep gene by changing the spatial distance between the gene and its two regulatory genes.
This approach seems to be consistent with our purpose. Then, under the guidance of our tutor, we did find two regulatory genes, CpxR and rpoE, related to the gene of dgep. We found that these two genes and dgep gene form a feedforward loop regulation relationship. Then, by adjusting the position of the gene in the genome, we can change the spatial distance between it and the two regulatory genes. In theory, we can change the expression of dgep gene. Therefore, we confirm our experimental technical scheme.
Because we try to use new technology, we need more technical experts. we interviewed Professor Xie of Huazhong University of Science and Technology, whose main research direction was microbiology and synthetic biology. He could help us more in our design from the perspective of synthetic biology. After the introduction of our projects.
Professor Xie gave affirmation to our design and creativity firstly. He suggested that the project is innovative and challenging for senior high. Then he pointed out that we needed to make the project design more integrated. Professor Xie helped us build up a deeper understanding of synthetic biology. He indicated that synthetic biology was more systematic, engineered and automatic than genetic engineering which modified targets on gene level and metabolic engineering which was on metabolic pathway.
Professor Xie pointed out that if we want to consider improving the heat resistance of engineered bacteria, it is important to set a suitable temperature gradient. Because the method we use is relatively advanced technology, there is a certain risk in the experiment. We can consider inviting experts of mathematical modeling to support our project to see whether the results of modeling meet our experimental expectations, and on this basis, we can draw up the scheme of experimental temperature gradient by consulting relevant literature.
In addition, Professor Xie pointed out that if we want to screen out high-quality strains, the setting of control strains is very important. It is not enough for us to set a single negative control strain, and it is also necessary to set up a positive control strain that has not been treated. Based on this theory, MG1655 is added to the design of our project_ Construction of MG1655_LCP strain.
To understand whether our project has more practical application, we visited Mr. Wang, an expert in BGI and microbial metagenomics. Mr. Wang and his colleagues told us that as the largest gene sequencing company in the world, BGI will undertake many projects from different institutions and enterprises, so they have some understanding of many applied projects. After we talked to them about our project and plan, they affirmed it and shared two problems that were hard to solve. I believe that once our project is successful, we will give great technical support in these two directions.
The first is sand fixation in the desert. In desert areas, in order to prevent the sandstorm, there is a kind of sticky protein to fix the sand particles together. However, the high temperature environment in these tropical areas will make the engineering bacteria unable to work well. Therefore, if the heat-resistant engineering strains can be transformed, it will be a very important basic contribution to the implementation of some projects in tropical areas.
In addition, in the process of organic fertilizer production by biological fermentation, the activity of some enzymes will increase with the increase of temperature, and the thermostable strain can also be used in these aspects. In addition, not only for the laboratory, but also for the production and fermentation process to improve the anti-impurity performance of the application.
Photo: field investigation and project presentation of BGI
Based on the above-mentioned public surveys and interviews, we have received feedback from the public and the experts. They all have positive feedback on improving the heat resistance of microorganisms. These positive feedbacks and generous donations from the general public show their support and encourage us that our project is good for everyone and does benefit the public.
In addition, our questionnaire shows that the public has a certain understanding of synthetic biology, which may be due to the publicity and promotion of our iGEM competition in the past two years. Therefore, we decided to design the popular science knowledge of synthetic biology more carefully.
In addition, we have revised our project design from the survey results and the opinions and suggestions of experts
1. Professor He's interview and BGI's on-site visit let us know more about the application direction of our project, which can not only be applied to the microbial fermentation industry, but also reduce the energy consumption cost. It can also be used in the production and fermentation process to improve the anti-impurity performance. In addition, there are important contributions to the implementation and application of some projects in tropical areas, such as the implementation and application of iGEM environment track.
2. During the interview with Professor Xie, he proposed to us the modeling and the design of positive control strains MG1655_LCP, and revised the direction of our project design, making our project more in line with academic norms.
3. Professor Ma provided us with the way of changing the space distance to adjust the expression of heat-resistant gene degp, which made us successfully explore a new path. In this way, the future iGEM team can not only use the various components we created to carry out its experimental operation, but also solve some problems from the perspective of gene space distance based on our ideas, so as to add more new vitality to the research in the field of synthetic biology.
