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RNAi

Period one,transformation, colony PCR, plasmid extraction, plasmid PCR, and infection experiments attempted

5.22-5.23

Xinxu Long,Ruoyu Chen——Transformation of RNAi plasmids; Inoculation of gray mold fungus onto PDA agar plates.

5.24-5.27

Xinxu Long,Ruoyu Chen——Plasmid extraction; Colony PCR of HT115(DE3) and subsequent gel electrophoresis analysis.

5.28-5.29

Xinxu Long,Xiaocheng Feng——Gray mold fungus infection on tomato fruits (inoculation with fungal agar plugs, inoculation with fungal spore suspension); Inoculation of gray mold fungus.

5.30

Xinxu Long——Plasmid extraction; Plasmid PCR and gel electrophoresis analysis of HT115(DE3).

6.1

Xinxu Long,Xiaocheng Feng——Gray mold fungus infection on tomato fruits (inoculation with circular filter paper soaked with fungal spore suspension); Scratch inoculation of gray mold fungus on agar plates for sporulation induction.

6.5-6.7

Xinxu Long,Xiaocheng Feng——Transformation of RNAi plasmids; Plasmid extraction; Colony PCR and gel electrophoresis analysis of HT115(DE3); Plasmid PCR and gel electrophoresis analysis of HT115(DE3).

7.5

Xinxu Long,Xiaocheng Feng——Gray mold fungus infection on tomato fruits and detached leaves, with four different infection methods attempted: agar plug inoculation, glass paper with gray mold, fungal spore suspension, circular filter paper soaked with fungal spore suspension.

Period two,shRNA extraction, infection experiment attempts

7.11-7.18

Ruoyu Chen,Xinxu Long,Xiaocheng Feng——Inoculation of gray mold fungus onto PDA agar plates; Shake culture, induction of Escherichia coli with IPTG, extraction of various shRNAs using Trizol method and completion of nucleic acid gel electrophoresis.

7.19-7.20

Xinxu Long,Xiaocheng Feng——Inoculation of gray mold fungus onto PDA agar plates; Shake culture, induction of Escherichia coli with IPTG, extraction of various shRNAs using Trizol method and completion of nucleic acid gel electrophoresis.

7.21

Xinxu Long,Xiaocheng Feng——Bulk extraction of shRNA and completion of nucleic acid gel electrophoresis; Gray mold fungus infection on tomato fruits and detached leaves (inoculation with agar plugs); Inoculation of gray mold fungus onto PDA agar plates.

7.22-7.24

Xinxu Long,Xiaocheng Feng——Bulk extraction of shRNA and completion of nucleic acid gel electrophoresis.

7.25

Xinxu Long,Xiaocheng Feng——After spraying with naked shRNA or CPP-coated shRNA, tomato fruits were inoculated with gray mold fungus using agar plugs; Transformation of GFP plasmid; Inoculation of gray mold fungus onto PDA agar plates.

7.26-7.28

Xinxu Long,Xiaocheng Feng——Bulk extraction of shRNA and completion of nucleic acid gel electrophoresis; Scratch inoculation of gray mold fungus on agar plates for sporulation induction; Inoculation of gray mold fungus onto PDA agar plates.

7.29

Xinxu Long,Xiaocheng Feng——Gray mold fungus infection on tomato fruits (inoculation with fungal spore suspension); Inoculation of gray mold fungus onto PDA agar plates.

7.30

Xinxu Long,Xiaocheng Feng——Bulk extraction of shRNA and completion of nucleic acid gel electrophoresis.

8.1

Xinxu Long,Xiaocheng Feng——Attempted to obtain gray mold spores and prepare spore suspension; Observed spore morphology under a microscope; Tried rapid sporulation method; Simulated natural gray mold infection on tomato fruits using a 5x4 egg carton.

8.2

Xinxu Long,Xiaocheng Feng——Gray mold fungus infection on tomato fruits (inoculation with fungal spore suspension and mycelial suspension); Bulk extraction of shRNA and completion of nucleic acid gel electrophoresis.

8.3

Xinxu Long,Xiaocheng Feng——Gray mold fungus infection on tomato fruits (inoculation with fungal spore suspension); Bulk extraction of shRNA and completion of nucleic acid gel electrophoresis.

8.4

Xinxu Long,Xiaocheng Feng——Inoculation of gray mold fungus onto PDA agar plates; Bulk extraction of shRNA and completion of nucleic acid gel electrophoresis.

8.5

Xinxu Long,Xiaocheng Feng——Gray mold fungus infection on tomato fruits (inoculation with fungal spore suspension); Bulk extraction of shRNA and completion of nucleic acid gel electrophoresis.

8.8

Xiaocheng Feng——Inoculation of gray mold fungus onto PDA agar plates; Bulk extraction of shRNA and completion of nucleic acid gel electrophoresis.

Period three,application of naked shRNA

8.10-8.11

Xiaocheng Feng,Jun Huang,Yutong Chen——After spraying with naked shRNA, tomato fruits were inoculated with small agar plugs of gray mold fungus.

8.12-8.14

Xiaocheng Feng,Jun Huang——Processing of lesion area data; Inoculation of gray mold fungus onto PDA agar plates.

8.15-8.16

Xiaocheng Feng,Jun Huang——After spraying naked shRNA, tomato fruits were inoculated with small agar plugs of gray mold fungus.
Xiaocheng Feng——Bulk extraction of shRNA and completion of nucleic acid gel electrophoresis.

8.17

Xiaocheng Feng——Bulk extraction of shRNA and completion of nucleic acid gel electrophoresis.
Yutong Chen——Inoculation of gray mold fungus onto PDA agar plates.

8.18

Xiaocheng Feng,Jun Huang——qPCR tomato sample collection.

8.19

Jun Huang——Inoculation of gray mold fungus onto PDA agar plates.
Xiaocheng Feng,Jun Huang,Yutong Chen——qPCR tomato sample collection; Bulk extraction of shRNA and completion of nucleic acid gel electrophoresis.
Xiaocheng Feng——Scratch inoculation of gray mold fungus on agar plates for sporulation induction; Microscopic observation of spore morphology.

Period four,application of shRNA coated with CPP (Cell Penetrating Peptide)

8.23

Xiaocheng Feng,Jun Huang——After spraying with shRNA coated with CPP (Cell Penetrating Peptide), tomato fruits were inoculated with small agar plugs of gray mold fungus.
Ruoyu Chen——Bulk extraction of shRNA.

8.24

Xiaocheng Feng,Jun Huang——After spraying with shRNA coated with CPP (Cell Penetrating Peptide), tomato fruits were inoculated with small agar plugs of gray mold fungus.
Yutong Chen——Inoculation of gray mold fungus onto PDA agar plates; Bulk extraction of shRNA.

8.26-8.27

Jiayi Huang——qPCR tomato sample collection.
Jun Huang——Inoculation of gray mold fungus onto PDA agar plates.

8.28

Xiaocheng Feng,Jun Huang,Xinxu Long——After spraying with shRNA coated with Cell Penetrating Peptide (CPP), tomato fruits were inoculated with small agar plugs of gray mold fungus.
Yutong Chen——Inoculation of gray mold fungus onto PDA agar plates.

8.29-9.2

Xiaocheng Feng,Xinxu Long——Sample collection of tomato samples treated with naked RNA; RNA extraction from tomato samples; Reverse transcription of RNA into cDNA; Primer selection; qRT-PCR (quantitative reverse transcription PCR).

9.4

Xiaocheng Feng,Xinxu Long——qRT-PCR (naked RNA treatment).

9.5

Xiaocheng Feng,Xinxu Long——Bulk extraction of shRNA.

9.6-9.14

Xiaocheng Feng,Xinxu Long——RNA extraction from tomato samples (RNA treatment with CPP coating); Reverse transcription of RNA into cDNA; qRT-PCR.

9.15-9.16

Xiaocheng Feng,Xinxu Long——After spraying with shRNA coated with CPP (Cell Penetrating Peptide), tomato fruits were inoculated with small agar plugs of gray mold fungus.

Period five,Validation of concatenated shRNA.

9.17

Xiaocheng Feng,Xinxu Long——RNA extraction from tomato samples (treated with concatenated RNA coated with CPP); Reverse transcription of RNA into cDNA; qRT-PCR; Repeat experiment - Spraying with naked shRNA followed by inoculation of tomato fruits with small agar plugs of gray mold fungus.

9.18-9.19

Xiaocheng Feng,Xinxu Long——qPCR of concatenated RNA.

9.20-9.28

Xiaocheng Feng,Xinxu Long——For continuous qPCR (shRNA stability test) infection experiments; Sample RNA, cDNA reverse transcription and qPT-PCR for continuous qPCR and bare shRNA were extracted.

5.18-5.27

Yutong Chen——Construction of PehA Plasmid

5.31-6.4

Ruoyu Chen,Tangkun Zhu——Transformation of PehA plasmid and inoculation of transformed BL21 (DE3) onto agar plates.

6.5-7.15

Yutong Chen,Tangkun Zhu——Plasmid extraction for PCR and enzyme digestion, followed by gel electrophoresis verification.

7.16-7.30

Yutong Chen,Tangkun Zhu——IPTG induction, extraction of PehA protein, protein concentration determination using the BCA method, protein electrophoresis using SDS-PAGE, and Western blot verification of PehA expression.

7.24-7.31

Yutong Chen,Tangkun Zhu,Yang Yang——Two methods, protoplast accumulation and extracellular alkalinization, were used to identify the immune response induced in leaves after flg22 application.
Ruoyu Chen——Construction of BvEP plasmid.

7.31-8.1

Ruoyu Chen,Tangkun Zhu——Activation of BL21 (DE3) containing the flg22 plasmid and inoculation onto agar plates.

8.2

Yutong Chen——Extraction of flg22 plasmid for PCR and enzyme digestion, followed by gel electrophoresis verification.

8.3-8.20

Yutong Chen,Tangkun Zhu——IPTG induction, extraction of flg22 protein, protein concentration determination using the BCA method, protein electrophoresis using SDS-PAGE, and Western blot validation of flg22 expression.

8.4-9.24

Yutong Chen,Tangkun Zhu,Yang Yang——DAB staining method for identifying the ROS burst in leaves induced by flg22 application.

8.9

Ruoyu Chen,Tangkun Zhu——Transformation of BvEP plasmid and inoculation of the transformed BL21 (DE3) onto agar plates.

8.10-8.20

Yutong Chen,Tangkun Zhu——IPTG induction, extraction of BvEP protein, protein concentration determination using the BCA method, protein electrophoresis using SDS-PAGE, and Western blot validation of BvEP expression.

8.21

Yutong Chen——Optimization of IPTG induction conditions for the expression of flg22 and BvEP.

8.22-9.24

Yutong Chen,Yang Yang——New IPTG induction, extraction of flg22 and BvEP proteins, protein concentration determination using the BCA method, protein electrophoresis using SDS-PAGE, and Western blot validation of flg22 and BVEP expression.

9.21

Yutong Chen,Tangkun Zhu,Yang Yang——Identification of ROS burst in leaves after application of PehA, flg22, and BvEP using luminescent staining method.

10.2

All experiments have concluded.

8.15

Tangkun Zhu, Ruoyu Chen——Summarize the current situation of pesticide use, including pesticide abuse and the use of RNAi biopesticides. Gets the inspiration of making the product into a laundry detergent beads from our PI. We our initial idea is to make our product into a bead.

8.25

Tangkun Zhu——Learns how the laundry detergent beads produce, finding the raw material for bead production.

9.10

Xinxu Long, Tangkun Zhu, Ruoyu Chen——We interview Mr. Xiang and Mr. Zhang from the Testing Center of Shenzhen University to discuss what kind of electron microscopy can be used to observe our biological samples.

9.13

Tangkun Zhu——Designs our product shapes, content and accessories. We decided to use PVA (polyvinyl alcohol) film as the material for our packaging.

9.14

Xinxu Long, Tangkun Zhu, Ruoyu Chen——After learning and comparing the use rules and observation objects of different electron microscopy, we decide to use scanning electron microscopy (SEM) to observe our RNA-CPP complex at the microscopic scale. We designed CPP, RNA and RNA-CPP complexes with different concentrations and observed them under SEM.

9.21

Xinxu Long, Tangkun Zhu, Ruoyu Chen——Use scanning electron microscopy (SEM) to observe our RNA-CPP complex at the microscopic scale. We use SEM for further observation and determine the structure of our sample under SEM.

9.24

Tangkun Zhu——Finds the most suitable solvent contained in our complex biopesticide products.

9.26-10.5

Tangkun Zhu——Designs the experiment of product verification and carry out the related test, and carry out the product production.

Dry lab

7.11-8.5

Jun Huang——Building an RNA interference model.

Jun Huang——Building an epidemiological model. The model guides how the product is applied.

8.5-9.3

Jun Huang——Helped with tomato infestation tests in the lab.

Jun Huang——Analyzing the data from the experiment.Data were analyzed to advance the experiment.

9.10-9.20

Jun Huang——validating the model against the data produced by the experiment.

Hardware

5.1-6.31

Zhonghao Sun——Conception of shRNA production by in vitro transcription.
Zhonghao Sun—— Preliminary design of hardware functions and structure.

7.1-7.20

Zhonghao Sun——Model selection of hardware components and microcontrollers.
Zhonghao Sun——3D Modeling of the mini reaction container with 3ds MAX.

7.21-8.31

Zhonghao Sun ——3D Printing of the mini reaction container.
Zhonghao Sun ——Assembly of various components.
Zhonghao Sun ——Writing and debugging of microcontroller programs.

9.1-10.2

Zhonghao Sun ——Attempt of producing shRNA using the built hardware.
Zhonghao Sun ——Verify the shRNA produced by the hardware.

Human practice

March

3.11

All members——Shenzhen University Affiliated Experimental High School visited the iGEM lab for learning and exploration, where we introduced synthetic biology to them.

3.20

Zuxin Chen,Xinyi Hao——Haijixing Agricultural Market Research: After learning about the threat of gray mold disease to tomatoes, we initiated a brainstorming session to develop a preliminary plan.

April

4.2

All members——Online exchange meeting with China Agricultural University to share our respective project brainstorms and provide suggestions to each other.

4.22

All members——Participated in the Deep Blue Charity Sale, where iGEM teams not only raised funds for public welfare projects but also contributed to the dissemination and promotion of synthetic biology knowledge. This helped promote innovation, education, and the spirit of public service.

4.23

All members——Innovation and Entrepreneurship Carnival, where entrepreneurs shared their startup stories and experiences, providing valuable advice for our future entrepreneurial plans.

May

5.13

Zuxin Chen,Xinyi Hao——Interview with Mr. Li Xiaojie, Deputy Director of the Technology Department at the Genomics Institute. He emphasized the importance of adopting a "prevention is better than cure" plant protection philosophy and highlighted the significance of focusing on product stability, delivery efficiency, and effectiveness.

5.21

All members——South China Exchange Conference, which brought together teams from the South China region to build a bridge for interactive displays, collaborative exchanges, and mutual learning, creating a conducive atmosphere for exploring synthetic biology together.

5.31

Zuxin Chen,Xinyi Hao——Interview with Professor Zhang Yongmin from Shenzhen University, who believes that plant vaccines play a crucial role in sustainable agriculture by effectively reducing the use of pesticides on plants.

July

7.11

All members——CCIC Conference, which brought together teams from different regions to facilitate interactive displays, collaborative exchanges, and mutual learning.

7.11

All members——We had a discussion with the team from Zhejiang University, exchanging ideas and discussing the project, while providing each other with suggestions.

7.16

All members——High school teams visited our lab for tours and exchanges, followed by project presentations and discussions.

7.16

All members——iGEM team from Macau University exchanged ideas, discussing projects and planning the first iGEM Greater Bay Area Synthetic Biology Industry-Academia-Research Forum.

7.20

Zuxin Chen,Xinyi Hao——The Shenzhen Agricultural Technology Promotion Center conducted an interview and exchange with Dr. Li Yonghong, Chief Researcher at the Shenzhen Agricultural Promotion Center, and Dr. Jin Man from the Chinese Academy of Sciences. They provided us with a broader perspective and targeted solutions.

7.22

Zuxin Chen,Xinyi Hao——We conducted interviews with agricultural markets and supermarkets, which will help us better meet market demands and consumer expectations with our products.

August

8.10

Zuxin Chen,Xinyi Hao,Ruoyu Chen,Tangkun Zhu,Jiayi Huang——We visited the Huizhou Boluo County Agricultural Technology Demonstration Farm to understand the usage of pesticides, the promotion methods for new products, and the promotional responsibilities undertaken by the department.

8.11

Zuxin Chen,Xinyi Hao,Ruoyu Chen,Tangkun Zhu,Jiayi Huang——Director Yin from the Pesticide Residue Testing Center of Boluo County Agricultural Bureau highly praised our biopesticides, emphasizing the importance of gaining recognition from farmers and ensuring effective pesticide performance.

8.11

Zuxin Chen,Xinyi Hao,Ruoyu Chen,Tangkun Zhu,Jiayi Huang——Interviewed Senior Engineer Wang Rifang and Teacher Li Li from the Agricultural Comprehensive Service Center of Boluo County Agricultural and Rural Service Center, discussing major tomato diseases and related issues.

8.12

Zuxin Chen,Xinyi Hao,Ruoyu Chen,Tangkun Zhu,Jiayi Huang——Visited Guangdong Hongke Testing Technology Co., Ltd., consulting them on specific pesticide residue testing procedures and touring their laboratory.

8.15

Zuxin Chen,Xinyi Hao,Ruoyu Chen,Tangkun Zhu,Jiayi Huang——We conducted an interview with Technician Hong at Huizhou BoshangJiaonong Ecological Agriculture Development Co., Ltd. Here, we gained insights into tomato cultivation and his unique approach to farming.

8.19

Zuxin Chen,Xinyi Hao,Honglin Wei,Yang Yang,Jiayi Huang——First iGEM Greater Bay Area Synthetic Biology Industry-Academia-Research Forum, exploring new ways of collaboration among iGEM teams and emphasizing the collaboration between academia and industry.

8.20

Zuxin Chen,Xinyi Hao,Ruoyu Chen,Erhuan Guan——University City Parent-Child Summer Camp, organizing a series of carefully planned activities to ensure meaningful experiences for the participants.

September

9.3

All members——2023 iGEM Intercollegiate Synthetic Biology Science Popularization Conference, we provided detailed explanations of our project to participating students and conducted science popularization on the safety of biopesticides.

9.10

All members——A series of Shenzhen University lectures aimed at gradually guiding college students to deepen their understanding of this field.

9.23

Zuxin Chen,Xinyi Hao,Yang Yang,Jiayi Huang——We visited the Dacheng Xin Qianxi Tomato Industry Base for an interview and to conduct trials using our products.

Art desigining

3.4-3.9

Jiayi Huang,Erhuan Guan——Making original gifts and designing game cards related to synthetic biology for students from Affiliated High School of Shenzhen University.

4.8-4.22

Jiayi Huang,Erhuan Guan——Designing materials for Shenlan Campus Sales and annual exposition for students'entrepreneuring and businessing.

4.23-5.1

Erhuan Guan——Designing team uniforms.

5.3-5.21

Jiayi Huang,Erhuan Guan——Designing the 7th Southern China Regional Meeting related materials.

7.5-7.7

Jiayi Huang,Erhuan Guan——Designing poster of our project for CCiC(Conference of China iGEMer Community).

7.10-7.20

Jiayi Huang—— Drawing emoji pics.

8.1-8.9

Jiayi Huang,Erhuan Guan——designing painting albums for education.

8.10-8.16

Jiayi Huang——Recording the human practice in huizhou .

8.28-9.14

Jiayi Huang——Completing promotion video.

9.15-10.12

Jiayi Huang,Erhuan Guan——Shooting the presentation video and designing the wiki.

Wiki Develop

6.20-7.15

Honglin Wei——Familiarize with the official iGEM wiki editing operations.

7.15-8.20

Honglin Wei—— learning HTML, CSS, and JavaScript.

8.20-9.15

Honglin Wei——Build the wiki framework for our SZU iGEM 2023.

9.15-10.12

Honglin Wei——Complete the home page and gradually complete other necessary pages.

Thanks
A big thank you to Ling Rongsong, Wu Yingfeng, Shen Yunfeng from Zhejiang University, and others for their help. I am extremely grateful to the other members of SZU 2023 iGEM team and our close collaboration, which allowed us to successfully complete the creation of this wiki!