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首页> 外文期刊>Marine biology >Single-step nested multiplex PCR to differentiate between various bivalve larvae
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Single-step nested multiplex PCR to differentiate between various bivalve larvae

机译：单步巢式多重PCR可以区分不同的双壳幼虫

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A nested multiplex polymerase chain reaction (PCR) assay has been developed that allows the discrimination between six bivalve larvae common to Danish coastal waters (Cerastoderma edule, Macoma balthica, Mytilus edulis, Spisula subtruncata, Ensis americanus and members of the order Myoida). This assay involves the simultaneous use of a pair of general universally targeted 18S rRNA gene primers, five specific 18S rRNA gene targeted oligonucleotide primers internal (nested) to the universal primer pair and one species-specific primer that is not nested (Mya). The specificity of each primer was evaluated in silico, empirically, and verified further by sequencing of amplification products from single larvae collected from plankton samples. Identification of individually isolated bivalve larvae from plankton samples was based on the size of the PCR product produced by the specific primers after visualisation by agarose gel electrophoresis. Preliminary studies indicated that this method was suitable for use with freshly collected and preserved larvae, and is therefore suitable for field application.
机译：已开发出一种嵌套的多重聚合酶链反应（PCR）分析方法，该方法可区分丹麦沿海水域常见的六个双壳类幼虫（Cerastoderma edule，Macoma balthica，Mytilus edulis，Spisula subtruncata，Ensis americanus和Myoida目成员）。该测定法涉及同时使用一对通用通用靶向18S rRNA基因引物，五个通用（通用）引物内部（嵌套）的18S rRNA基因靶向寡核苷酸引物和一个未嵌套的物种特异性引物（Mya）。每种引物的特异性都通过计算机进行了经验评估，并通过对从浮游生物样品中收集的单个幼虫的扩增产物进行测序来进一步验证。从浮游生物样品中分离出的双壳幼虫的鉴定是基于琼脂糖凝胶电泳显像后，由特定引物产生的PCR产物的大小。初步研究表明，该方法适用于新鲜收集和保存的幼虫，因此适合现场应用。

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