微流控富集条形码（MEBarcoding）：一种高通量植物 DNA 条形码的新方法。
Microfluidic Enrichment Barcoding (MEBarcoding): a new method for high throughput plant DNA barcoding.
机构信息
Botanical Research Institute of Texas, Fort Worth, Texas, 76107-3400, USA.
Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases (NIAID), NIH, Bethesda, MD, 20892, USA.
出版信息
Sci Rep. 2020 May 26;10(1):8701. doi: 10.1038/s41598-020-64919-z.
DNA barcoding is a valuable tool to support species identification with broad applications from traditional taxonomy, ecology, forensics, food analysis, and environmental science. We introduce Microfluidic Enrichment Barcoding (MEBarcoding) for plant DNA Barcoding, a cost-effective method for high-throughput DNA barcoding. MEBarcoding uses the Fluidigm Access Array to simultaneously amplify targeted regions for 48 DNA samples and hundreds of PCR primer pairs (producing up to 23,040 PCR products) during a single thermal cycling protocol. As a proof of concept, we developed a microfluidic PCR workflow using the Fluidigm Access Array and Illumina MiSeq. We tested 96 samples for each of the four primary DNA barcode loci in plants: rbcL, matK, trnH-psbA, and ITS. This workflow was used to build a reference library for 78 families and 96 genera from all major plant lineages - many currently lacking in public databases. Our results show that this technique is an efficient alternative to traditional PCR and Sanger sequencing to generate large amounts of plant DNA barcodes and build more comprehensive barcode databases.
DNA 条形码是一种支持物种鉴定的有价值的工具，具有广泛的应用，包括传统分类学、生态学、法医学、食品分析和环境科学。我们引入了用于植物 DNA 条形码的微流控富集条形码（MEBarcoding），这是一种具有成本效益的高通量 DNA 条形码方法。MEBarcoding 使用 Fluidigm Access Array 同时扩增 48 个 DNA 样本的靶向区域和数百个 PCR 引物对（在单个热循环方案中产生多达 23040 个 PCR 产物）。作为概念验证，我们使用 Fluidigm Access Array 和 Illumina MiSeq 开发了一种微流控 PCR 工作流程。我们对四个主要植物条形码基因座（rbcL、matK、trnH-psbA 和 ITS）的每个基因座测试了 96 个样本。该工作流程用于构建来自所有主要植物谱系的 78 个科和 96 个属的参考文库 - 许多目前在公共数据库中缺失。我们的结果表明，该技术是一种替代传统 PCR 和 Sanger 测序的有效方法，可用于生成大量植物 DNA 条形码并构建更全面的条形码数据库。
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