蜱媒病毒研究中蜱的解剖和离体器官培养

Current Protocols in Microbiology Pub Date : 2020-10-08 DOI:10.1002/cpmc.118

Jeffrey M. Grabowski, Ryan Kissinger

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引用次数: 6

摘要

蜱传病毒每年在全世界造成数千例疾病。针对许多蜱传病毒的具体对策还没有市售。关于蜱-病毒相互作用的了解很少，增加这方面的知识可以导致开发对策的潜在目标。蜱体外器官培养的病毒感染可以提供一种鉴定易感细胞类型组织感染的方法。此外，这些器官培养可用于功能基因组研究，以查明参与病毒生命周期的蜱特异性基因。这里提供了逐步建立基本蜱器官培养结合病毒感染和/或功能基因组研究的程序。这些程序可适用于将来用于表征其他蜱传病原体感染以及蜱特有的生物过程。©2020 Wiley期刊有限公司基本方案1:用明胶泡沫底板加载96孔板基本方案2:对未喂食的雌/雄肩胛骨蜱进行多器官分步无菌解剖基本方案3:对喂食的雌肩胛骨蜱进行分步无菌解剖以去除唾液腺基本方案4:蜱器官培养物的代谢活力分析基本方案5:蜱器官培养物的病毒感染基本方案6:蜱器官培养的功能性RNA干扰分析

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Ixodid Tick Dissection and Tick Ex Vivo Organ Cultures for Tick-Borne Virus Research

Tick-borne viruses cause thousands of cases of disease worldwide every year. Specific countermeasures to many tick-borne viruses are not commercially available. Very little is known regarding tick-virus interactions and increasing this knowledge can lead to potential targets for countermeasure development. Virus infection of ex vivo organ cultures from ticks can provide an approach to identify susceptible cell types of tissue to infection. Additionally, these organ cultures can be used for functional genomic studies to pinpoint tick-specific genes involved in the virus lifecycle. Provided here are step-by-step procedures to set up basic tick organ cultures in combination with virus infection and/or functional genomic studies. These procedures can be adapted for future use to characterize other tick-borne pathogen infections as well as tick-specific biological processes. © 2020 Wiley Periodicals LLC.

Basic Protocol 1: Loading 96-well plates with gelfoam substrate

Basic Protocol 2: Step-by-step aseptic dissection of unfed female/male Ixodes scapularis ticks for multiple organs

Basic Protocol 3: Step-by-step aseptic dissection of fed female Ixodes scapularis ticks to remove salivary glands

Basic Protocol 4: Metabolic viability analyses of tick organ cultures

Basic Protocol 5: Virus infection of tick organ cultures

Basic Protocol 6: Functional RNA interference analyses using tick organ cultures

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来源期刊

Current Protocols in Microbiology Immunology and Microbiology-Parasitology

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期刊介绍： Current Protocols in Microbiology provides detailed, step-by-step instructions for analyzing bacteria, animal and plant viruses, fungi, protozoans and other microbes. It offers updated coverage of emerging technologies and concepts, such as biofilms, quorum sensing and quantitative PCR, as well as proteomic and genomic methods. It is the first comprehensive source of high-quality microbiology protocols that reflects and incorporates the new mandates and capabilities of this robust and rapidly evolving discipline.