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首页> 外文期刊>Applied and Environmental Microbiology >Optimization of Terminal-Restriction Fragment Length Polymorphism Analysis for Complex Marine Bacterioplankton Communities and Comparison with Denaturing Gradient Gel Electrophoresis
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Optimization of Terminal-Restriction Fragment Length Polymorphism Analysis for Complex Marine Bacterioplankton Communities and Comparison with Denaturing Gradient Gel Electrophoresis

机译：复杂海洋浮游生物群落末端限制性片段长度多态性分析的优化及其与变性梯度凝胶电泳的比较

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摘要

The potential of terminal-restriction fragment length polymorphism (T-RFLP) and the detection of operational taxonomic units (OTUs) by capillary electrophoresis (CE) to characterize marine bacterioplankton communities was compared with that of denaturing gradient gel electrophoresis (DGGE). A protocol has been developed to optimize the separation and detection of OTUs between 20 and 1,632 bp by using CE and laser-induced fluorescence detection. Additionally, we compared T-RFLP fingerprinting to DGGE optimized for detection of less abundant OTUs. Similar results were obtained with both fingerprinting techniques, although the T-RFLP approach and CE detection of OTUs was more sensitive, as indicated by the higher number of OTUs detected. We tested the T-RFLP fingerprinting technique on complex marine bacterial communities by using the 16S rRNA gene and 16S rRNA as templates for PCR. Samples from the Northern and Middle Adriatic Sea and from the South and North Aegean Sea were compared. Distinct clusters were identifiable for different sampling sites. Thus, this technique is useful for rapid evaluation of the biogeographical distribution and relationships of bacterioplankton communities.
机译：将末端限制性片段长度多态性（T-RFLP）和通过毛细管电泳（CE）检测海洋生物浮游生物群落特征性操作分类单位（OTU）的潜力与变性梯度凝胶电泳（DGGE）的潜力进行了比较。已经开发出一种协议，通过使用CE和激光诱导的荧光检测来优化20到1,632 bp之间OTU的分离和检测。此外，我们将T-RFLP指纹图谱与为检测次丰度OTU而优化的DGGE进行了比较。尽管T-RFLP方法和OTU的CE检测更为灵敏，但两种指纹技术均获得了相似的结果，这表明检测到的OTU数量更多。我们使用16S rRNA基因和16S rRNA作为PCR模板，在复杂的海洋细菌群落上测试了T-RFLP指纹技术。比较了北部和中亚得里亚海以及南部和北爱琴海的样品。对于不同的采样点，可以识别出不同的簇。因此，该技术可用于快速评估浮游生物群落的生物地理分布和关系。

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