Development of real-time PCR technique for the estimation of population density of Pythium intermedium in forest soils.
Development of real-time PCR technique for the estimation of population density of Pythium intermedium in forest soils.
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DOI：
10.1016/j.micres.2009.11.010
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发表时间：
2010-10
影响因子：
6.7
通讯作者：
Mingzhu Li;Masako Senda;T. Komatsu;H. Suga;K. Kageyama
中科院分区：
文献类型：
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作者：
Mingzhu Li;Masako Senda;T. Komatsu;H. Suga;K. Kageyama
在寒温带森林土壤的碳循环中，中皮在土壤中起着重要的作用。本研究利用物种特异性引物，建立了一种快速、精确、有效的实时PCR技术，用于估算土壤中介苗的种群密度。用常规PCR和实时PCR验证特异性。测定检测限（灵敏度），用SYBR Green II荧光染料生成扩增标准曲线。采用一种新的DNA提取方法和实时PCR引物，建立了一种快速、准确的测定日本高山森林土壤中间苗的方法。采用土壤电镀法和实时荧光定量PCR技术，对森林土壤中间苗的分布进行了研究。这项新技术将为研究python物种在森林和农业生态系统中的作用提供有用的工具，并可应用于实际应用。
Pythium intermedium is known to play an important role in the carbon cycling of cool-temperate forest soils. In this study, a fast, precise and effective real-time PCR technique for estimating the population densities of P. intermedium from soils was developed using species-specific primers. Specificity was confirmed both with conventional PCR and real-time PCR. The detection limit (sensitivity) was determined and amplification standard curves were generated using SYBR Green II fluorescent dye. A rapid and accurate assay for quantification of P. intermedium in Takayama forest soils of Japan was developed using a combination of a new DNA extraction method and PCR primers were developed for real-time PCR. And the distribution of P. intermedium in forest soil was investigated with both soil plating method and the developed real-time PCR technique. This new technique will be a useful tool and can be applied to practical use for studying the role of Pythium species in forest and agricultural ecosystems.