Biomass measurement by flow cytometry during solid‐state fermentation of basidiomycetes
Biomass measurement by flow cytometry during solid‐state fermentation of basidiomycetes
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担子菌固态发酵过程中流式细胞术的生物量测量
DOI：
10.1002/cyto.a.22592
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Bley Th
中科院分区：
文献类型：
--
作者：
Steudler S;Böhmer U;Weber J;Bley Th
固态发酵（SSF）是一种稳健的工艺，非常适合现场培养担子菌，产生用于处理木质纤维素的酶。可靠的生物量定量方法对于真菌生长动力学分析至关重要。然而，在 SSF 期间不可能直接测定生物量，因为真菌会生长到基质中并将其用作营养源。这就需要使用间接方法，这些方法要么非常费力且耗时，要么只能提供特定生长时期的生物量测量。在这里，我们描述了一种基于流式细胞术计数真菌细胞核的 SSF 期间真菌生物量快速测定新方法的开发和优化。真菌生物量在有机基质上生长，其浓度通过细胞破碎后从真菌菌丝中分离出细胞核，用 SYTOX®Green 染色，然后使用流式细胞仪计数来测量。建立了将样品的干生物量与其细胞核浓度联系起来的校准曲线。测试了多种缓冲液和破坏方法。将获得的结果与使用麦角甾醇测定方法测定的值进行比较，麦角甾醇测定方法是 SSF 期间真菌生物量测量的经典技术。我们的新方法可用于测量各种不同规模的真菌生物量，从 15 mL 培养物到工作体积为 10 L 的实验室反应器（由医疗技术和生物技术研究中心 (fzmb GmbH) 开发）。 © 2014 国际细胞计数促进会
Solid‐state fermentation (SSF) is a robust process that is well suited to the on‐site cultivation of basidiomycetes that produce enzymes for the treatment of lignocellulosics. Reliable methods for biomass quantification are essential for the analysis of fungal growth kinetics. However, direct biomass determination is not possible during SSF because the fungi grow into the substrate and use it as a nutrient source. This necessitates the use of indirect methods that are either very laborious and time consuming or can only provide biomass measurements during certain growth periods. Here, we describe the development and optimization of a new rapid method for fungal biomass determination during SSF that is based on counting fungal nuclei by flow cytometry. Fungal biomass was grown on an organic substrate and its concentration was measured by isolating the nuclei from the fungal hyphae after cell disruption, staining them with SYTOX®Green, and then counting them using a flow cytometer. A calibration curve relating the dry biomass of the samples to their concentrations of nuclei was established. Multiple buffers and disruption methods were tested. The results obtained were compared with values determined using the method of ergosterol determination, a classical technique for fungal biomass measurement during SSF. Our new approach can be used to measure fungal biomass on a range of different scales, from 15 mL cultures to a laboratory reactor with a working volume of 10 L (developed by the Research Center for Medical Technology and Biotechnology (fzmb GmbH)). © 2014 International Society for Advancement of Cytometry
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DOI：
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发表时间：
2008
期刊：
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DOI：
10.1002/cyto.990190203
发表时间：
1995-02-01
期刊：
CYTOMETRY
影响因子：
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作者：
DOLEZEL, J;GOHDE, W
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影响因子：
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作者：
J. Weber;V. Georgiev;A. Pavlov;T. Bley
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T. Bley
影响因子：
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A. W. West;D. J. Ross;J. C. Cowling