DNA mini-barcoding

  • 文章类型: Journal Article
    蘑菇中毒是全球食源性疾病和相关死亡的重要原因。Amanita蘑菇经常引起这种中毒;然而,由于无法获得新鲜和完整的样品,因此识别这些有毒物种具有挑战性。通常需要分析残留物,呕吐物,或胃提取物以获得DNA序列,用于鉴定导致食物中毒的物种。这通常证明获得可用的DNA序列具有挑战性,所述可用的DNA序列可以使用常规分子生物学技术进行分析。因此,这项研究旨在开发一种DNA迷你条形码方法,用于鉴定天牛物种。在对Amanita蘑菇中DNA迷你条形码的通用引物进行评估和优化之后,我们发现内部转录间隔区(ITS)基因序列引物ITS-a是鉴定天牛物种最合适的DNA条形码引物。随后扩增并测序了43个Amanita样品。对获得的序列进行了种内和种间遗传距离分析,并构建了系统发育树。结果表明,所设计的引物在天牛样品中具有很强的普适性,可以准确鉴定出长度为290bp的目的基因片段。值得注意的是,DNA迷你条形码准确识别了43个天牛样本,证明与常规DNA条形码的高度一致性。此外,它有效地从消化样品中鉴定出DNA。总之,这种DNA迷你条形码是一种有前途的工具,用于检测意外摄入有毒的鹅膏菌。它可以用作最佳条形码,用于在天牛引起的蘑菇中毒事件中进行物种识别和可追溯性。关键点:•开发用于无新鲜样品的天牛物种鉴定的DNA迷你条形码方法。•ITS-a引物集经优化以在天牛样品中实现稳健的通用性。•迷你条形码适用于在蘑菇中毒情况下筛选有毒蘑菇物种。
    Mushroom poisoning contributes significantly to global foodborne diseases and related fatalities. Amanita mushrooms frequently cause such poisonings; however, identifying these toxic species is challenging due to the unavailability of fresh and intact samples. It is often necessary to analyze residues, vomitus, or stomach extracts to obtain DNA sequences for the identification of species responsible for causing food poisoning. This usually proves challenging to obtain usable DNA sequences that can be analyzed using conventional molecular biology techniques. Therefore, this study aimed to develop a DNA mini-barcoding method for the identification of Amanita species. Following the evaluation and optimization of universal primers for DNA mini-barcoding in Amanita mushrooms, we found that the internal transcribed spacer (ITS) gene sequence primer ITS-a was the most suitable DNA barcode primer for identifying Amanita species. Forty-three Amanita samples were subsequently amplified and sequenced. The sequences obtained were analyzed for intra- and inter-species genetic distances, and a phylogenetic tree was constructed. The findings indicated that the designed primers had strong universality among the Amanita samples and could accurately identify the target gene fragment with a length of 290 bp. Notably, the DNA mini-barcode accurately identified the 43 Amanita samples, demonstrating high consistency with the conventional DNA barcode. Furthermore, it effectively identified DNA from digested samples. In summary, this DNA mini-barcode is a promising tool for detecting accidental ingestion of toxic Amanita mushrooms. It may be used as an optimal barcode for species identification and traceability in events of Amanita-induced mushroom poisoning. KEY POINTS: • Development of a DNA mini-barcoding method for Amanita species identification without fresh samples. • The ITS-a primer set was optimized for robust universality in Amanita samples. • The mini-barcode is suitable for screening toxic mushroom species in mushroom poisoning cases.
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  • 文章类型: Journal Article
    如今,鉴于食品认证在质量和安全方面的相关性,它越来越需要。海鲜市场受到错误标签和欺诈性替换/掺假的严重影响,特别是加工食品,如罐头食品,由于形态特征的丧失。这项研究旨在开发基于DNA的新测定方法,以鉴定新鲜的鲭鱼(Scomberspp。)和商业产品。在5SrRNA基因和非转录间隔区(NTS)上重新设计了一个新的引物对,鉴定适用于加工的商业产品的物种鉴定的DNA迷你条形码区。此外,提供快速和低成本的分析,开发了一种基于重组酶聚合酶扩增(RPA)的新方法,用于鉴定新鲜的“Sgombro”(Sccomberscombrus)和“LanzardooOcchione”(Sccomberjaponicus和Sccombercolias),再加上最昂贵的物种(Scomberscombrus)识别的侧向流动可视化。这种创新的便携式检测方法在海鲜市场的供应链可追溯性方面具有巨大潜力。
    UNASSIGNED:
    UNASSIGNED:在线版本包含补充材料,可在10.1007/s12161-022-02429-6获得。
    Nowadays, food authentication is more and more required given its relevance in terms of quality and safety. The seafood market is heavily affected by mislabelling and fraudulent substitutions/adulterations, especially for processed food products such as canned food items, due to the loss of morphological features. This study aims to develop new assays based on DNA to identify fresh mackerel (Scomber spp.) and commercial products. A new primer pair was de novo designed on the 5S rRNA gene and non-transcribed spacer (NTS), identifying a DNA mini-barcoding region suitable for species identification of processed commercial products. Moreover, to offer a fast and low-cost analysis, a new assay based on recombinase polymerase amplification (RPA) was developed for the identification of fresh \'Sgombro\' (Scomber scombrus) and \'Lanzardo o Occhione\' (Scomber japonicus and Scomber colias), coupled with the lateral flow visualisation for the most expensive species (Scomber scombrus) identification. This innovative portable assay has great potential for supply chain traceability in the seafood market.
    UNASSIGNED:
    UNASSIGNED: The online version contains supplementary material available at 10.1007/s12161-022-02429-6.
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  • 文章类型: Journal Article
    UNASSIGNED: The dry body of the Tokay Gecko (Gekko gecko) is the source of a valuable traditional Chinese medicine, it is therefore listed as a Class II protected animal species in China. Due to increasing market demand and a declining supply of the species, a considerable number of adulterants have emerged in the market. Thus, it is necessary to establish an accurate and rapid method of identification for distinguishing G. gecko from its adulterants and for separating it from highly processed products.
    UNASSIGNED: A total of 274 COI sequences were analyzed by using MEGA 5.0 software. Several specific primers were designed to amplify mini-barcode regions and identify G. gecko from its counterfeits and products.
    UNASSIGNED: 274 COI sequences of G. gecko and 15 adulterants species were analyzed. G. gecko could be distinguished from its adulterants through BLAST analysis, intra- and inter-specific distance analyses, and an NJ tree based on COI sequences. Two pairs of specific primers designed for this study, COISF2/COISR2 and COISF3/COISR3, amplified 200- and 133-bp fragments of the COI region, respectively, both of which were suitable for the identification of G. gecko and its adulterants. Furthermore, COISF3/COISR3 detected G. gecko in 15 batches of products.
    UNASSIGNED: Therefore, the specific DNA mini-barcoding method developed here may be a powerful tool for the identification of G. gecko and counterfeits, and may also be used to distinguish G. gecko from its highly processed by-products.
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  • 文章类型: Journal Article
    In recent years, the demand for natural herbal products (NHP) has increased; however, the quality of these products is difficult to confirm due to the lack of a comprehensive quality control system. Traditional methods are not effective in detecting processed ingredients. DNA barcoding is an established technique that has been used for more than 10 years. This technique uses short standard sequences (generally 200-600 bp) to identify species. While a complete DNA barcode is difficult to obtain from NHP due to DNA degradation, mini-barcoding is a complementary tool to identify species in NHP. DNA mini-barcoding uses smaller DNA segments for polymerase chain reaction amplification and can be applied to identify species rapidly. The present review summarizes the development and application of DNA mini-barcodes over recent years and discusses the limitations of this technique. This review also compares mini-barcoding and meta-barcoding, a technique using universal polymerase chain reaction primers to simultaneously amplify multiple DNA barcodes and identify many species in a single environmental sample. Additionally, other detection methods that can be combined with mini-barcodes, such as nucleotide signatures, high-resolution DNA melting analysis, and gold nanoparticles, are discussed. DNA mini-barcoding can fill the gaps left by other methods in the field of herbal molecular identification.
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