University of Chinese Academy of Sciences 27 articles published in JoVE Bioengineering An Integrated Micro-Device System for Coral Growth and Monitoring Jinglun Zhao1, Tao Yuan2, Hui Huang2, Xiaolin Lu1 1State Key Laboratory of Digital Medical Engineering, School of Biological Science & Medical Engineering, Southeast University, 2Key Laboratory of Tropical Marine Bio-resources and Ecology, Guangdong Provincial Key Laboratory of Applied Marine Biology, South China Sea Institute of Oceanology, University of Chinese Academy of Sciences This protocol describes the development of a modular controllable micro-device system that can be applied for the long-term culturing and monitoring of sea corals. Environment Collection of Alfalfa Root Exudates to Study the Impact of Di(2-ethylhexyl) Phthalate on Metabolite Production Wenjie Ren1,2, Rui Zhao1, Ying Teng1, Yongming Luo1 1Key Laboratory of Soil Environment and Pollution Remediation, Institute of Soil Science, Chinese Academy of Sciences, 2College of Resources and Environment, University of Chinese Academy of Sciences The secretion of root exudates is usually an external detoxification strategy for plants under stress conditions. This protocol describes how to assess the impact of xenobiotics on alfalfa via nontargeted metabolomic analysis. Biology A Precise and Quantifiable Method for Collecting Hemolymph from Small Arthropods Qing Liu1,3, Lili Zhang1,2, Rongxiang Fang1,2,3, Yan Huo1,2 1State Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese Academy of Sciences, 2CAS Center for Excellence in Biotic Interactions, University of Chinese Academy of Sciences, 3University of Chinese Academy of Sciences We describe a method to collect quantifiable hemolymph efficiently from small arthropods for subsequent analysis. Developmental Biology An Efficient Clearing Protocol for the Study of Seed Development in Tomato (Solanum lycopersicum L.) Yixuan Feng1,2, Tai Wang1,2,3, Lingtong Liu1 1Key Laboratory of Plant Molecular Physiology, Institute of Botany, Chinese Academy of Sciences, 2College of Life Science, University of Chinese Academy of Sciences, 3Innovative Academy of Seed Design, Chinese Academy of Sciences The tomato seed is an important model for studying genetics and developmental biology during plant reproduction. This protocol is useful for clearing tomato seeds at different developmental stages to observe the finer embryonic structure. Bioengineering Cellular Affinity of Particle-Stabilized Emulsion to Boost Antigen Internalization Fengqiang Cao*1,2,3, Yali Ming*3,4, Weixiang Gao3,4, Jia Ge3, Kenji Ogino1,2 1Graduate School of Bio-Applications and Systems Engineering, Tokyo University of Agriculture and Technology, 2Institute of Global Innovation Research, Tokyo University of Agriculture and Technology, 3State Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, 4University of Chinese Academy of Sciences To rationally design efficient adjuvants, we developed poly-lactic-co-glycolic acid nanoparticle-stabilized Pickering emulsion (PNPE). The PNPE possessed unique softness and a hydrophobic interface for potent cellular contact and offered high-content antigen loading, improving the cellular affinity of the delivery system to antigen-presenting cells and inducing efficient internalization of antigens. Biology Mosquito-Associated Virus Isolation from Field-Collected Mosquitoes Doudou Huang*1, Haixia Ma*1, Lu Zhao1,2, Xiaoyu Wang1,2, Yi Huang1,2, Fei Wang1, Zhiming Yuan1,2, Han Xia1,2 1Key Laboratory of Special Pathogens and Biosafety, Wuhan Institute of Virology, Chinese Academy of Sciences, 2University of Chinese Academy of Sciences Numerous novel virus-like sequences have been found in mosquitoes due to the extensive use of sequencing technologies. We provide an effective procedure for isolating and amplifying viruses using vertebrate and mosquito cell lines, which might serve as the basis for future studies on mosquito-associated viruses, including mosquito-borne and mosquito-specific viruses. Immunology and Infection Experimental Viral Infection in Adult Mosquitoes by Oral Feeding and Microinjection Fei Wang*1, Cihan Yang*1,2, Shunlong Wang1,2, Qun Wu1,2, Christabel Ochieng1,2, Zhiming Yuan1,2, Han Xia1,2 1Key Laboratory of Special Pathogens and Biosafety, Wuhan Institute of Virology, Chinese Academy of Sciences, 2University of Chinese Academy of Sciences This methodology, which included oral feeding and intrathoracic injection infection, could effectively assess the influence of midgut and/or salivary gland barriers on arbovirus infection. Engineering Rapid Repetition Rate Fluctuation Measurement of Soliton Crystals in a Microresonator Peng Xie*1,2, Xinyu Wang*1,2, Weiqiang Wang1,2, Wenfu Zhang1,2, Zhizhou Lu1,2, Yang Wang1,2, Wei Zhao1,2 1State Key Laboratory of Transient Optics and Photonics, Xi'an Institute of Optics and Precision Mechanics (XIOPM), Chinese Academy of Sciences (CAS), 2University of Chinese Academy of Sciences Here, we present a protocol to generate soliton crystals in a butterfly-packaged micro-ring resonator using a thermal tuned method. Further, the repetition rate fluctuations of a soliton crystal with a single vacancy are measured using a delayed self-heterodyne method. Immunology and Infection Preparation of Pseudo-Typed H5 Avian Influenza Viruses with Calcium Phosphate Transfection Method and Measurement of Antibody Neutralizing Activity Xiaoyan Chang*1, Keru Zhou*1, Yanguang Liu1,2, Lian Duan3, Lichun Zhang2, Guoliang Zhang3, Haikun Wang4, Guiqin Wang1 1Nanjing Advanced Academy of Life and Health, 2Institute of Animal Bio-technology, Jilin Academy of Agricultural Sciences, 3National Clinical Research Center for Infectious Diseases, Shenzhen Third People’s Hospital, Southern University of Science and Technology, 4CAS Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, University of Chinese Academy of Sciences Here we describe a protocol for pseudovirus packaging and the measurement of antibody neutralizing activity. Environment Ecotoxicological Effects of Microplastics on Bird Embryo Development by Hatching without Eggshell Liyi Wang1,2, Nana Xue1,2, Wenfeng Li2, Rehemanjiang Wufuer1, Daoyong Zhang1,3 1Xinjiang Key Laboratory of Environmental Pollution and Bioremediation, Xinjiang Institute of Ecology and Geography, Chinese Academy of Sciences, 2University of Chinese Academy of Sciences, 3Key Laboratory of Microbial Technology for Industrial Pollution Control of Zhejiang Province, College of Environment, Zhejiang University of Technology This paper introduces a method of hatching without using an eggshell for toxicological studies of particle pollutants such as microplastics. Biology Two-Step Reverse Transcription Droplet Digital PCR Protocols for SARS-CoV-2 Detection and Quantification Raphael Nyaruaba1,2,3, Xiaohong Li1, Caroline Mwaliko2,3,4, Changchang Li1, Matilu Mwau5, Nelson Odiwour1,2,3, Elishiba Muturi1,2,3, Caroline Muema1,2,3, Junhua Li1, Junping Yu1, Hongping Wei1 1Key Laboratory of Special Pathogens and Biosafety, Center for Biosafety Mega-Science, Wuhan Institute of Virology, Chinese Academy of Sciences, 2International College, University of Chinese Academy of Sciences, 3Sino-Africa Joint Research Center, 4CAS Key Laboratory of Molecular Virology & Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, 5Center for Infectious and Parasitic Diseases Control Research, Kenya Medical Research Institute This work summarizes steps on developing different assays for SARS-CoV-2 detection using a two color ddPCR system. The steps are elaborate and notes have been included on how to improve the assays and experiment performance. These assays may be used for multiple SARS-CoV-2 RT-ddPCR applications. Immunology and Infection Stability and Structure of Bat Major Histocompatibility Complex Class I with Heterologous β2-Microglobulin Di Zhang*1,2, Kefang Liu*3,4, Dan Lu*2,5, Pengyan Wang1,2, Qingxu Zhang1,2, Peipei Liu2, Yingze Zhao2, Yan Chai4, Jianxin Lyu1, Jianxun Qi4, William J. Liu1,2 1School of Laboratory Medicine and Life Sciences, Wenzhou Medical University, 2NHC Key Laboratory of Biosafety, National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, 3Faculty of Health Sciences, University of Macau, 4CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, 5Savaid Medical School, University of Chinese Academy of Sciences The protocol describes experimental methods to obtain stable major histocompatibility complex (MHC) class I through potential β2-microglobulin (β2m) substitutions from different species. The structural comparison of MHC I stabilized by homologous and heterologous β2m were investigated. Engineering Improving the Combustion Performance of a Hybrid Rocket Engine using a Novel Fuel Grain with a Nested Helical Structure Zezhong Wang1,2, Xin Lin1, Fei Li1, Zelin Zhang1, Xilong Yu1,2 1Institute of Mechanics, Chinese Academy of Sciences, 2School of Engineering Science, University of Chinese Academy of Sciences A technique utilizing a solid fuel grain with a novel nested helical structure to improve the combustion performance of a hybrid rocket engine is presented. Immunology and Infection Evaluation of T Follicular Helper Cells and Germinal Center Response During Influenza A Virus Infection in Mice Meng Wang*1, Yuanyuan Huang*1, Wangpeng Gu2,3, Haikun Wang1 1CAS Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, University of Chinese Academy of Sciences, 2School of Life Science, University of Science and Technology of China, 3State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences This paper describes protocols of evaluating Tfh and GC B response in mouse model of influenza virus infection. Engineering Graphene-Assisted Quasi-van der Waals Epitaxy of AlN Film on Nano-Patterned Sapphire Substrate for Ultraviolet Light Emitting Diodes Xiang Zhang*1,2, Zhaolong Chen*3, Hongliang Chang1,2, Jianchang Yan1,2, Shenyuan Yang2,4, Junxi Wang1,2, Peng Gao5, Tongbo Wei1,2 1State Key Laboratory of Solid-State Lighting, Institute of Semiconductors, Chinese Academy of Sciences, 2Center of Materials Science and Optoelectronics Engineering, University of Chinese Academy of Science, 3Center for Nanochemistry (CNC), Beijing National Laboratory for Molecular Science, College of Chemistry and Molecular Engineering, Peking University, 4State Key Laboratory of Superlattices and Microstructures, Institute of Semiconductors, Chinese Academy of Sciences, 5Electron Microscopy Laboratory, School of Physics, Peking University A protocol for graphene-assisted growth of high-quality AlN films on nano-patterned sapphire substrate is presented. Genetics Tomato Root Transformation Followed by Inoculation with Ralstonia Solanacearum for Straightforward Genetic Analysis of Bacterial Wilt Disease Rafael J. L. Morcillo1, Achen Zhao1,2, María I. Tamayo-Navarrete3, José M. García-Garrido3, Alberto P. Macho1 1Shanghai Center for Plant Stress Biology, CAS Center for Excellence in Molecular Plant Sciences, Shanghai Institutes of Biological Sciences, Chinese Academy of Sciences, 2University of Chinese Academy of Sciences, 3Department of Soil Microbiology and Symbiotic Systems, Estación Experimental del Zaidín (CSIC) Here, we present a versatile method for tomato root transformation followed by inoculation with Ralstonia solanacearum to perform straightforward genetic analysis for the study of bacterial wilt disease. Engineering Thermocapillary Convection Space Experiment on the SJ-10 Recoverable Satellite Li Duan*1,2, Yongli Yin*3, Jia Wang*1, Qi Kang1,2, Di Wu1, Huan Jiang1, Pu Zhang1, Liang Hu1 1National Microgravity Laboratory, Institute of Mechanics, Chinese Academy of Sciences, 2School of Engineering Sciences, University of Chinese Academy of Sciences, 3China Astronaut Research and Training Center A protocol for the space payload design, the space experiment on thermocapillary convection, and analyses of experimental data and images are presented in this paper. Chemistry Isotopic Effect in Double Proton Transfer Process of Porphycene Investigated by Enhanced QM/MM Method Zhihui Tu1,2, Jian Yin3, Liangxu Xie4 1State Key Laboratory of Structural Chemistry, Fujian Institute of Research on the Structure of Matter, Chinese Academy of Sciences, 2University of Chinese Academy of Sciences, 3XtalPi Inc. (Shenzhen Jingtai Technology Co., Ltd.), 4Institute of Bioinformatics and Medical Engineering, School of Electrical and Information Engineering, Jiangsu University of Technology A protocol that uses enhanced QM/MM method to investigate the isotopic effect on the double proton transfer process in porphycene is presented here. Engineering Quantitative Analysis of Vacuum Induction Melting by Laser-induced Breakdown Spectroscopy Tianzhuo Zhao1,2,3, Xin Li1,2, Qixiu Zhong1,2, Hong Xiao1,3, Shuzhen Nie1,3, Fuqiang Lian1,3, Sining Sun4, Zhongwei Fan1,2,3 1The Academy of Opto-electronics, Chinese Academy of Sciences, 2University of Chinese Academy of Sciences, 3National Engineering Research Center for DPSSL, 4Beijing GK Laser Technology Co., Ltd. During vacuum induction melting, laser-induced breakdown spectroscopy is used to perform real-time quantitative analysis of the main-ingredient elements of a molten alloy. Chemistry Separation of Uranium and Thorium for 230Th-U Dating of Submarine Hydrothermal Sulfides Lisheng Wang1,2,3, Xuefeng Wang1,2, Jun Ye4, Zhibang Ma1,2, Weifang Yang5, Jule Xiao1,2 1Key Laboratory of Cenozoic Geology and Environment, Institute of Geology and Geophysics, Chinese Academy of Sciences, 2Institutions of Earth Science, Chinese Academy of Sciences, 3University of Chinese Academy of Sciences, 4First Institute of Oceanography, State Oceanic Administration, 5Key Laboratory of Submarine Geosciences, Second Institute of Oceanography, State Oceanic Administration The protocol describes a method to purify and separate the U and Th nuclide in submarine hydrothermal sulfide sample with Fe co-precipitation and extraction chromatography for 230Th-U disequilibrium dating. Behavior A Conflict Model of Reward-seeking Behavior in Male Rats Shaofei Jiang1,2, Yue Zhang1,2, Xigeng Zheng1,2, Haoshuang Luo1,2, Zhengkui Liu1,2, Yunjing Bai1,2 1CAS Key Laboratory of Mental Health, Institute of Psychology, Chinese Academy of Sciences, 2Department of Psychology, University of Chinese Academy of Sciences This conflict model is used to measure the impairment of inhibitory control after exposure to addictive drugs, or other factors that may influence inhibitory control. A sexual stimulus and an aversive obstacle are concurrently presented, thus male rats have to conquer the obstacle to approach the sexual reward. Biology Visualizing the Interaction Between the Qdot-labeled Protein and Site-specifically Modified λ DNA at the Single Molecule Level Huijun Xue*1,2, Zhengyan Zhan*1, Kaining Zhang1,2, Yu Vincent Fu1,2 1State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, 2Savaid Medical School, University of Chinese Academy of Sciences Here, we present a protocol to study DNA-protein interactions by total internal reflection fluorescence microscopy (TIRFM) using a site-specifically modified λ DNA substrate and a Quantum-dot labeled protein. Behavior Combining Quantitative Food-intake Assays and Forcibly Activating Neurons to Study Appetite in Drosophila Lifen Jiang*1,2, Yinpeng Zhan*2,3, Yan Zhu2,3 1School of Life Science, University of Science and Technology of China, 2State key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, 3University of Chinese Academy of Sciences Quantitative food-intake assays with dyed food provide a robust and high-throughput means to evaluate feeding motivation. Combining the food consumption assay with thermogenetic and optogenetic screens is a powerful approach to investigate the neural circuits underlying appetite in adult Drosophila melanogaster. Developmental Biology A RANKL-based Osteoclast Culture Assay of Mouse Bone Marrow to Investigate the Role of mTORC1 in Osteoclast Formation Qinggang Dai*1, Yujiao Han*2, Furong Xie*1, Xuhui Ma3, Zhan Xu2, Xiao Liu1, Weiguo Zou2, Jun Wang1 1Department of Pediatric Dentistry, Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai Key Laboratory of Stomatology & Shanghai Research Institute of Stomatology, National Clinical Research Center of Stomatology, 2State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, University of Chinese Academy of Sciences, 3Department of Oral and Maxillofacial-Head and Neck Oncology, Ninth People's Hospital, Shanghai Jiaotong University School of Medicine This manuscript describes a protocol to isolate and culture osteoclasts in vitro from mouse bone marrow, and to study the role of the mammalian/mechanistic target of rapamycin complex 1 in osteoclast formation. Genetics CRISPR/Cas9-mediated Targeted Integration In Vivo Using a Homology-mediated End Joining-based Strategy Xuan Yao1,2, Xing Wang1,2, Junlai Liu2,3,4, Linyu Shi1, Pengyu Huang3, Hui Yang1 1Institute of Neuroscience, State Key Laboratory of Neuroscience, Key Laboratory of Primate Neurobiology, CAS Center for Excellence in Brain Science and Intelligence Technology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 2College of Life Sciences, University of Chinese Academy of Sciences, 3School of Life Science and Technology, Shanghai Tech University, 4Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences The clustered regularly interspaced short palindromic repeats/CRISPR associated protein 9 (CRISPR/Cas9) system provides a promising tool for genetic engineering, and opens up the possibility of targeted integration of transgenes. We describe a homology-mediated end joining (HMEJ)-based strategy for efficient DNA targeted integration in vivo and targeted gene therapies using CRISPR/Cas9. Developmental Biology Application of Aorta-gonad-mesonephros Explant Culture System in Developmental Hematopoiesis Junhua Lv1,2, Feng Liu1,2 1State Key Laboratory of Membrane Biology, Institute of Zoology, Chinese Academy of Sciences, 2University of Chinese Academy of Sciences This protocol describes using cultured Aorta-Gonad-Mesonephros for expression analyses, colony-forming units in the culture and spleen, and long-term reconstitution to determine the effect of regulatory factors and signaling pathways on hematopoietic stem cell development. This has been demonstrated as an effective system for studying hematopoietic stem cell biology and function. Engineering The Modular Design and Production of an Intelligent Robot Based on a Closed-Loop Control Strategy Libo Zhang1, Junjie Zhu2, Hao Ren2, Dongdong Liu3, Dan Meng4, Yanjun Wu1, Tiejian Luo5 1Institute of Software, Chinese Academy of Sciences, 2Hunan University, 3Fuyang Normal University, 4Anhui Normal University, 5University of Chinese Academy of Sciences We present a protocol on modular design and production of intelligent robots to help scientific and technical workers design intelligent robots with special production tasks based on personal needs and individualized design.