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应用聚合酶链反应-限制性片段长度多态性和蛋白质组学分析鉴定北方长额虾(Pandalus borealis)。

Species identification of the Northern shrimp (Pandalus borealis) by polymerase chain reaction-restriction fragment length polymorphism and proteomic analysis.

机构信息

Department of Analytical Chemistry, Nutrition, and Food Science, School of Veterinary Sciences, University of Santiago de Compostela, E-27002 Lugo, Spain.

出版信息

Anal Biochem. 2012 Feb 1;421(1):56-67. doi: 10.1016/j.ab.2011.10.029. Epub 2011 Oct 22.

Abstract

Genomic and proteomic techniques for species identification of meat and seafood products are being widely used. In this study, a genomic approach was used to differentiate Pandalus borealis (the Northern shrimp), which belongs to the superfamily Pandaloidea, from 30 crustaceans consisting of 19 commercially relevant prawns/shrimps species that belong to the superfamily Penaeoidea, which include the families Penaeidae and Solenoceridae, and 11 other crustacean species, including prawns, shrimps, lobsters, and crabs. For this purpose, a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method was designed based on the amplification of the 16S rRNA/tRNA(Val)/12S rRNA mitochondrial regions using the primers 16S-CruF and 16S-CruR. The 966-bp PCR products were produced and cleaved with the restriction enzymes AluI, TaqI, and HinfI, which provided species-specific restriction patterns. In addition, a proteomic approach, based on matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) and electrospray ionization-ion trap (ESI-IT) mass spectrometry, was used to identify and characterize new P. borealis-specific peptides that could be useful as potential markers of this species in protein-based detection methods. To our knowledge, this is the first time a molecular method has been successfully applied to identify a wide range of prawn and shrimp species, including P. borealis, for either whole individuals or processed products. However, validation of the methods proposed here is required by applying them to a larger sample of individuals from different populations and geographic origins in order to avoid mainly false-negative results.

摘要

基因组和蛋白质组技术广泛用于肉类和海鲜产品的物种鉴定。在这项研究中,采用基因组方法从属于对虾超科的 30 种甲壳类动物中区分北方长额虾(北方虾),这 30 种甲壳类动物包括 19 种商业相关的对虾/虾类,属于真虾超科,包括对虾科和蝉虾科,以及 11 种其他甲壳类动物,包括对虾、虾、龙虾和蟹。为此,使用引物 16S-CruF 和 16S-CruR 对 16S rRNA/tRNA(Val)/12S rRNA 线粒体区域进行扩增,设计了聚合酶链反应-限制性片段长度多态性 (PCR-RFLP) 方法。产生了 966-bp 的 PCR 产物,并使用限制酶 AluI、TaqI 和 HinfI 进行切割,提供了物种特异性的限制图谱。此外,还采用基于基质辅助激光解吸/电离飞行时间 (MALDI-TOF) 和电喷雾电离-离子阱 (ESI-IT) 质谱的蛋白质组学方法来鉴定和表征新的北方长额虾特异性肽,这些肽可能作为该物种在基于蛋白质的检测方法中的潜在标记物。据我们所知,这是首次成功应用分子方法来鉴定广泛的对虾和虾类物种,包括北方长额虾,无论是整个个体还是加工产品。然而,需要通过将这些方法应用于来自不同种群和地理起源的更大个体样本进行验证,以避免主要的假阴性结果。

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