Using rDNA ITS2 barcoding to identify kratom (Mitragyna speciosa) from the genus Mitragyna and Neolamarckia cadamba.

IF 3 3区生物学 Q2 BIOCHEMICAL RESEARCH METHODS

ELECTROPHORESIS Pub Date : 2024-07-11 DOI:10.1002/elps.202400003

Meng-Yi Chen, Yu-Ching Tu, Hsin-Yi Shyu, Ting-An Lin, Chun-Pai Juan, Fang-Chin Wu

{"title":"Using rDNA ITS2 barcoding to identify kratom (Mitragyna speciosa) from the genus Mitragyna and Neolamarckia cadamba.","authors":"Meng-Yi Chen, Yu-Ching Tu, Hsin-Yi Shyu, Ting-An Lin, Chun-Pai Juan, Fang-Chin Wu","doi":"10.1002/elps.202400003","DOIUrl":null,"url":null,"abstract":"<p><p>This study collected 80 samples of suspected kratom plant powder. A polymerase chain reaction sequence analysis was conducted using two sets of DNA barcode primers for plant ribosomal (r)DNA internal transcribed spacers (ITSs), namely, ITS3/ITS4 and ITS-p3/ITS-u4. Among the 80 samples, 40 were analyzed using the ITS3/ITS4 primer pair, and then DNA sequences were subjected to a National Center for Biotechnology Information-Basic Local Alignment Search Tool (NCBI-BLAST) comparison. Results showed that 29 samples had a 100% match (364/364) with Mitragyna speciosa (kratom), and 6 samples had a 99.73% match (363/364) with M. speciosa, whereas 5 samples had disordered and unreadable sequences. The 5 unreadable samples and an additional 40 suspected kratom samples were then analyzed using the ITS-p3/ITS-u4 primer pair, followed by an NCBI-BLAST comparison. Among these, 32 samples had a 100% match (404/404) with M. speciosa, and 11 samples had a 99.75% match (403/404) with M. speciosa. Among the samples with sequences matching M. speciosa, three distinct types were observed (no variance/404, 287M/404, and 287A/404). One sample had a 99.51% match (404/406) with Neolamarckia cadamba, and another sample had a sequencing length of 305 bp, with 25 positions showing mixed base pairs, indicating a mixture of different species. Analysis of the mixed base pair pattern suggested a possible mixture of M. speciosa and N. cadamba. Actually, M. speciosa and N. cadamba have very similar external morphologies. This indicates that the ITS-p3/ITS-u4 primer pair is effective in distinguishing mixtures of M. speciosa and N. cadamba and is thus more suitable than ITS3/ITS4 for identifying and analyzing samples of suspected kratom plant powder.</p>","PeriodicalId":11596,"journal":{"name":"ELECTROPHORESIS","volume":" ","pages":""},"PeriodicalIF":3.0000,"publicationDate":"2024-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"ELECTROPHORESIS","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1002/elps.202400003","RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}

引用次数: 0

Abstract

This study collected 80 samples of suspected kratom plant powder. A polymerase chain reaction sequence analysis was conducted using two sets of DNA barcode primers for plant ribosomal (r)DNA internal transcribed spacers (ITSs), namely, ITS3/ITS4 and ITS-p3/ITS-u4. Among the 80 samples, 40 were analyzed using the ITS3/ITS4 primer pair, and then DNA sequences were subjected to a National Center for Biotechnology Information-Basic Local Alignment Search Tool (NCBI-BLAST) comparison. Results showed that 29 samples had a 100% match (364/364) with Mitragyna speciosa (kratom), and 6 samples had a 99.73% match (363/364) with M. speciosa, whereas 5 samples had disordered and unreadable sequences. The 5 unreadable samples and an additional 40 suspected kratom samples were then analyzed using the ITS-p3/ITS-u4 primer pair, followed by an NCBI-BLAST comparison. Among these, 32 samples had a 100% match (404/404) with M. speciosa, and 11 samples had a 99.75% match (403/404) with M. speciosa. Among the samples with sequences matching M. speciosa, three distinct types were observed (no variance/404, 287M/404, and 287A/404). One sample had a 99.51% match (404/406) with Neolamarckia cadamba, and another sample had a sequencing length of 305 bp, with 25 positions showing mixed base pairs, indicating a mixture of different species. Analysis of the mixed base pair pattern suggested a possible mixture of M. speciosa and N. cadamba. Actually, M. speciosa and N. cadamba have very similar external morphologies. This indicates that the ITS-p3/ITS-u4 primer pair is effective in distinguishing mixtures of M. speciosa and N. cadamba and is thus more suitable than ITS3/ITS4 for identifying and analyzing samples of suspected kratom plant powder.

查看原文本刊更多论文

利用 rDNA ITS2 条形码从 Mitragyna 和 Neolamarckia cadamba 属中识别桔梗（Mitragyna speciosa）。

本研究收集了 80 份疑似桔梗的植物粉末样本。研究人员使用两组针对植物核糖体（r）DNA 内部转录间隔（ITSs）的 DNA 条形码引物（即 ITS3/ITS4 和 ITS-p3/ITS-u4）进行聚合酶链式反应序列分析。在 80 个样本中，有 40 个样本使用 ITS3/ITS4 引物对进行了分析，然后对 DNA 序列进行了美国国家生物技术信息中心-基本局部比对搜索工具（NCBI-BLAST）比对。结果表明，29 个样本与桔梗（Mitragyna speciosa）的匹配度为 100%（364/364），6 个样本与桔梗的匹配度为 99.73%（363/364），而 5 个样本的序列混乱且无法读取。随后，使用 ITS-p3/ITS-u4 引物对这 5 个无法读取的样本和另外 40 个疑似桔梗样本进行了分析，并进行了 NCBI-BLAST 比对。其中，32 个样本与 M. speciosa 的匹配度为 100%（404/404），11 个样本与 M. speciosa 的匹配度为 99.75%（403/404）。在序列与 M. speciosa 匹配的样本中，观察到三种不同的类型（无差异/404、287M/404 和 287A/404）。其中一个样本与 Neolamarckia cadamba 的匹配度为 99.51%（404/406），另一个样本的测序长度为 305 bp，其中 25 个位置出现混合碱基对，表明混合了不同的物种。对混合碱基对模式的分析表明，M. speciosa 和 N. cadamba 可能是混合物。实际上，M. speciosa 和 N. cadamba 的外部形态非常相似。这表明 ITS-p3/ITS-u4 引物对能有效区分 M. speciosa 和 N. cadamba 的混合物，因此比 ITS3/ITS4 更适合鉴定和分析疑似桔梗植物粉末样本。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

求助全文

约1分钟内获得全文求助全文

来源期刊

ELECTROPHORESIS 生物-分析化学

CiteScore

6.30

自引率

13.80%

发文量

244

审稿时长

1.9 months

期刊介绍： ELECTROPHORESIS is an international journal that publishes original manuscripts on all aspects of electrophoresis, and liquid phase separations (e.g., HPLC, micro- and nano-LC, UHPLC, micro- and nano-fluidics, liquid-phase micro-extractions, etc.). Topics include new or improved analytical and preparative methods, sample preparation, development of theory, and innovative applications of electrophoretic and liquid phase separations methods in the study of nucleic acids, proteins, carbohydrates natural products, pharmaceuticals, food analysis, environmental species and other compounds of importance to the life sciences. Papers in the areas of microfluidics and proteomics, which are not limited to electrophoresis-based methods, will also be accepted for publication. Contributions focused on hyphenated and omics techniques are also of interest. Proteomics is within the scope, if related to its fundamentals and new technical approaches. Proteomics applications are only considered in particular cases. Papers describing the application of standard electrophoretic methods will not be considered. Papers on nanoanalysis intended for publication in ELECTROPHORESIS should focus on one or more of the following topics: • Nanoscale electrokinetics and phenomena related to electric double layer and/or confinement in nano-sized geometry • Single cell and subcellular analysis • Nanosensors and ultrasensitive detection aspects (e.g., involving quantum dots, "nanoelectrodes" or nanospray MS) • Nanoscale/nanopore DNA sequencing (next generation sequencing) • Micro- and nanoscale sample preparation • Nanoparticles and cells analyses by dielectrophoresis • Separation-based analysis using nanoparticles, nanotubes and nanowires.