Developing an SCAR and ITS reliable multiplex PCR-based assay for safflower adulterant detection in saffron samples
Developing an SCAR and ITS reliable multiplex PCR-based assay for safflower adulterant detection in saffron samples
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DOI：
10.1016/j.foodcont.2013.07.019
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发表时间：
2014-01-01
期刊：
影响因子：
6
通讯作者：
Bahar, Masoud
中科院分区：
文献类型：
--
作者：
Babaei, Saeid;Talebi, Majid;Bahar, Masoud
藏红花 (Crocus sativus L) 是国际贸易中最重要、最昂贵的药用香料产品之一，其设计或默认掺入红花雄蕊和玉米柱头，导致藏红花样品质量较差。本研究旨在开发特异性、灵敏且可重复的基于 PCR 的标记物，以检测贸易藏红花中的这些掺假物质。通过对真正的藏红花、红花和玉米进行比较 RAPD 分析，将随机引物 OPA-14、MG-11、MG-12 和 AJ-05 生成的 6 个推定 RAPD 标记鉴定为藏红花特异性标记。对这些特异性 RAPD 标记进行了克隆、测序并设计了六对 SCAR 引物。专门设计的引物能够以预期的大小扩增可重复的藏红花 DNA，而在玉米和红花 DNA 中则没有扩增。本研究还根据ITS序列设计了引物对，用于红花DNA的特异性扩增。 PCR反应还特异性扩增了红花基因组中ITS区域的613bp。进一步建立了多重 PCR 检测方法，以有效联合使用一些 SCAR 和 ITS 标记。这种新的分子方法的特点是技术快速、实用方便，适合分析大量样品。因此，基于简单快速PCR的分子方法可以作为鉴定藏红花掺假样品的有用辅助工具。本研究描述了一种基于 SCAR 和 ITS 标记的新型多重 PCR 检测的开发，用于快速分子检测藏红花中的替代品。 (C) 2013 Elsevier Ltd. 保留所有权利。
Saffron (Crocus sativus L), one of the most important and expensive medicinal spice products traded internationally, is subject to adulteration by design or default with safflower stamens and corn stigmas, leading to poor quality of saffron samples. The present study aims at the development of specific, sensitive and reproducible PCR-based markers to detect these adulterants in traded saffron. Six putative RAPD markers generated by random primers, OPA-14, MG-11, MG-12 and AJ-05, were identified as saffron specific by comparative RAPD analysis of genuine saffron, safflower and corn. These specific RAPD markers were cloned, sequenced and six pairs of SCAR primers were designed. Specific designed primers were able to amplify reproducible saffron DNA with expected sizes and no amplification in corn and safflower DNA. In this study, a primer pair was also designed based on ITS sequences for specific amplification of safflower DNA. PCR reactions were also specifically amplified 613 bp of ITS region in safflower genome. The multiplex PCR assays were further established for the joint use of some SCAR and ITS markers efficiently. The special feature of this new molecular method was technically rapid and convenient practically and suitable for analyzing large numbers of samples. Thus, the simple rapid PCR-based molecular method could be used as a helpful assistant tool for the identification of adulterant saffron samples. This study described the development of a new SCAR and ITS maker-based multiplex PCR assay for the rapid molecular detection of substitutes in saffron. (C) 2013 Elsevier Ltd. All rights reserved.