A next generation semiconductor based sequencing approach for the identification of meat species in DNA mixtures.
A next generation semiconductor based sequencing approach for the identification of meat species in DNA mixtures.
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DOI：
10.1371/journal.pone.0121701
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发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
Fontanesi L
中科院分区：
文献类型：
--
作者：
Bertolini F;Ghionda MC;D'Alessandro E;Geraci C;Chiofalo V;Fontanesi L
肉类和肉制品原产地物种的识别是预防和发现可能产生经济、道德和健康影响的欺诈行为的一个重要问题。在本文中，我们评估了下一代基于半导体的测序技术（Ion Torrent 个人基因组机）通过对从扩增 12S 和 16S rRNA 线粒体 DNA 基因的不同通用引物对获得的 PCR 产物进行测序，鉴定来自肉类物种（猪、马、牛、羊、兔、鸡、火鸡、雉鸡、鸭、鹅和鸽子）以及 DNA 混合物中的人类和大鼠 DNA 的潜力。产生了六个文库，包括分别从 13 个物种获得的 PCR 产物，或从包含来自所有物种或仅鸟类或仅哺乳动物物种的 DNA 的 DNA 混合物中获得的 PCR 产物，其摩尔浓度或猪和马 DNA 的比例为 1:10 或 1:50。测序共获得33,294,511个叫核苷酸，其中Q20为29,109,688个（87.43%），总共215,944个reads。使用不同的比对算法根据序列数据分配物种。通过桑格测序确认所获得的序列后计算出的错误率对于不同物种为0.0003至0.02。不同文库之间每个物种的读数数量的相关性对于哺乳动物物种较高（0.97），而对于鸟类物种较低（0.70）。 PCR 竞争限制了鸟类物种某些引物对的扩增和测序效率。使用从不同引物对获得的读数可以检测低水平的猪和马 DNA。对从不同通用 PCR 引物获得的产物进行测序可能是克服潜在扩增问题的有用策略。基于这些结果，Ion Torrent 技术可用于 DNA 混合物中肉类种类的鉴定。
The identification of the species of origin of meat and meat products is an important issue to prevent and detect frauds that might have economic, ethical and health implications. In this paper we evaluated the potential of the next generation semiconductor based sequencing technology (Ion Torrent Personal Genome Machine) for the identification of DNA from meat species (pig, horse, cattle, sheep, rabbit, chicken, turkey, pheasant, duck, goose and pigeon) as well as from human and rat in DNA mixtures through the sequencing of PCR products obtained from different couples of universal primers that amplify 12S and 16S rRNA mitochondrial DNA genes. Six libraries were produced including PCR products obtained separately from 13 species or from DNA mixtures containing DNA from all species or only avian or only mammalian species at equimolar concentration or at 1:10 or 1:50 ratios for pig and horse DNA. Sequencing obtained a total of 33,294,511 called nucleotides of which 29,109,688 with Q20 (87.43%) in a total of 215,944 reads. Different alignment algorithms were used to assign the species based on sequence data. Error rate calculated after confirmation of the obtained sequences by Sanger sequencing ranged from 0.0003 to 0.02 for the different species. Correlation about the number of reads per species between different libraries was high for mammalian species (0.97) and lower for avian species (0.70). PCR competition limited the efficiency of amplification and sequencing for avian species for some primer pairs. Detection of low level of pig and horse DNA was possible with reads obtained from different primer pairs. The sequencing of the products obtained from different universal PCR primers could be a useful strategy to overcome potential problems of amplification. Based on these results, the Ion Torrent technology can be applied for the identification of meat species in DNA mixtures.
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DOI：
10.1111/j.1471-8286.2006.01428.x
发表时间：
2006-12-01
期刊：
MOLECULAR ECOLOGY NOTES
影响因子：
--
作者：
Ivanova, Natalia V.;Dewaard, Jeremy R.;Hebert, Paul D. N.
通讯作者：
Hebert, Paul D. N.
影响因子：
4
作者：
Forde, Brian M.;O'Toole, Paul W.
通讯作者：
O'Toole, Paul W.
DOI：
10.1073/pnas.1308939110
发表时间：
2014-04-29
影响因子：
11.1
作者：
Flink, Linus Girdland;Allen, Richard;Larson, Greger
通讯作者：
Larson, Greger
影响因子：
4.1
作者：
Horreo, Jose L.;Ardura, Alba;Garcia-Vazquez, Eva
通讯作者：
Garcia-Vazquez, Eva
影响因子：
16
作者：
Heinz S;Benner C;Spann N;Bertolino E;Lin YC;Laslo P;Cheng JX;Murre C;Singh H;Glass CK
通讯作者：
Glass CK