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利用Cellena® portable microencapsulator将真菌包裹入400μm, 1,66%藻酸盐微胶囊（见图1），然后培养，最后用COPAS SELECT流式细胞术检测指标：颗粒大小(TOF), 光学密度 (EXT), 绿色自发荧光和红色自发荧光（见图2）
Genetic analysis of non-filamentous microorganisms is facilitated by the isolation of consistent, well‐defined colonies on solid media and the handling of individual cells by flow cytometry. In contrast, some filamentous fungi are hard to be analyzed using these procedures; in particular by flow cytometry. The combination of single spores microencapsulation and large particle flow cytometry is a possible alternative for the analysis of filamentous fungi. Microencapsulation allows the early detection of fungal growth by monitoring the development of hyphae from encapsulated individual spores. Mycelium proliferation inside the microcapsules can be detected using COPAS™ large particle flow cytometry.
Here we show the successful application of the Flow Focusing® technology to the microencapsulation of filamentous fungi in monodisperse alginate microspheres, using Aspergillus and Trychoderma as model systems. Using a Cellena® Flow Focusing
microencapsulator, we managed to produce monodisperse microparticles containing individual spores and to develop microcolonies of these fungi upon germination in the appropriate conditions. Proliferation inside the particles was monitored by microscopy and
large particle flow cytometry without requiring fluorescent labeling. Sterility was preserved during the microencapsulation procedure, preventing undesired contaminations. Conditional mutants were utilized to demonstrate the feasibility of the method. This procedure allows for the handling, screening and analysis of clonal colonies in liquid culture. Examples of applications are provided.
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