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首页> 外文期刊>Research in Microbiology >Digitized fluorescent RFLP analysis (fRFLP) as a universal method for comparing genomes of culturable dsDNA viruses: application to bacteriophages.
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Digitized fluorescent RFLP analysis (fRFLP) as a universal method for comparing genomes of culturable dsDNA viruses: application to bacteriophages.

机译：数字荧光RFLP分析（fRFLP）作为比较可培养dsDNA病毒基因组的通用方法：应用于噬菌体。

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摘要

Identification and classification of bacteriophages remains a cumbersome process even with the use of genotypic approaches, due to the lack of genes present in all phages. Restriction fragment length polymorphism analysis (RFLP) of the viral genome is a universal approach, but RFLP fingerprints obtained on agarose gels remain difficult to compare between laboratories. Here we describe the digitization of RFLP of viral genomes by amplification of all restriction fragments - after ligation of adapters - using primers complementary to the adapters only. Since one of the primers is fluorescently labelled, the restriction fragments become visible to a fluorescent capillary electrophoresis system (ABI310) and their lengths can be digitized immediately. The digitized fluorescent RFLP (fRFLP) fingerprint can be stored as an entry in a library. Dendrogram construction of the fRFLP fingerprints obtained for a total of 69 Caudovirales (tailed bacteriophages) showed that genomically and/or serologically closely related phages clustered, whereas host range was not completely in correspondence with genotype. fRFLP might be a tool for quickly establishing the relationship of newly isolated phages to previously isolated ones and for constructing an fRFLP library electronically accessible on the internet, to which fRFLP patterns of new phages can be compared.
机译：由于缺乏所有噬菌体中存在的基因，即使使用基因型方法，噬菌体的鉴定和分类仍然是一个繁琐的过程。病毒基因组的限制性片段长度多态性分析（RFLP）是一种通用方法，但是在琼脂糖凝胶上获得的RFLP指纹仍然很难在实验室之间进行比较。在这里，我们描述了通过仅在与衔接子互补的引物的情况下，通过扩增所有限制性片段（在衔接子连接后）来扩增病毒基因组RFLP的数字化。由于其中一种引物已被荧光标记，因此限制性片段对于荧光毛细管电泳系统（ABI310）变得可见，并且其长度可以立即数字化。可以将数字化的荧光RFLP（fRFLP）指纹作为条目存储在库中。共获得69个拟南芥（尾噬菌体）的fRFLP指纹的树状图构建显示，基因组和/或血清学上密切相关的噬菌体聚集在一起，而宿主范围并不完全与基因型相对应。 fRFLP可能是一种工具，可用于快速建立新分离的噬菌体与先前分离的噬菌体之间的关系，并构建可在互联网上以电子方式访问的fRFLP库，可以将新噬菌体的fRFLP模式与之进行比较。

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