基于鸟枪蛋白组学与质谱多反应监测技术的三文鱼物种鉴别研究

Authentication of salmon species based on shotgun proteomics and mass spectrometry multiple reaction monitoring techniques

中文关键词:  三文鱼  物种鉴别  鸟枪蛋白组学  多反应监测  特征肽段

英文关键词:salmon  species identification  shotgun proteomics  multiple reaction monitoring  specific peptide

中文摘要:

      目的 对大西洋鲑(Salmo salar)、大马哈鱼(Oncorhynchus keta)和虹鳟(Oncorhynchus mykiss)中的多肽进行分离鉴定, 筛选特征肽段, 建立不同种类三文鱼的物种鉴别方法。方法 以市场上常见的大西洋鲑、大马哈鱼和虹鳟3个物种为研究对象, 基于鸟枪蛋白组学分析流程, 样品经蛋白质提取、胰蛋白酶消化和超高效液相色谱-串联三重四极杆飞行时间质谱法(ultra performance liquid chromatography-tandem quadrupole time-of-flight mass spectrometry, UPLC-QTOF-MS)分离鉴定, 得到的总离子流图谱(total ion chromatogram, TIC)与UniProt蛋白质数据库对比分析, 筛选得到3个物种的特征肽段, 利用Skyline软件构建特征肽的多反应监测(multiple reaction monitoring, MRM)离子对, 然后经三重四极杆质谱仪进行鉴定。结果 最终共筛选出7条特征肽段, 包括大西洋鲑中的4条特征肽段, 大马哈鱼中的1条特征肽段, 虹鳟中的2条特征肽段。结论 基于鸟枪蛋白组学, 结合多反应监测技术, 筛选得到了大西洋鲑、大马哈鱼和虹鳟3个物种中的特征肽段, 成功建立了不同种类三文鱼的物种精准定性鉴别方法。

英文摘要:

      Objective To isolate and identify polypeptides from Salmo salar, Oncorhynchus keta and Oncorhynchus mykiss, screen the specific peptides, and establish the identification methods for different species of salmon. Methods Three common species of salmons, namely Atlantic salmon, Chum salmon and rainbow trout in the market were selected as the research objects, based on the shotgun proteomics protocol, the samples were separated and identified by ultra performance liquid chromatography-tandem quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) after protein extraction and trypsin digestion, the total ion chromatogram (TIC) obtained was compared with UniProt protein database to screen the specific peptides of 3 salmon species, the multiple reaction monitoring (MRM) ion pairs of the specific peptides were constructed by Skyline software, and then identified by triple quadrupole mass spectrometer. Results A total of 7 specific peptides including 4 specific peptides in Atlantic salmon, 1 in Chum salmon and 2 in rainbow trout were identified. Conclusion Based on shotgun proteomics, combined with multiple reaction monitoring techniques, the specific peptides of Atlantic salmon, Chum salmon and rainbow trout have been screened, and a precise qualitative identification method for different species of salmon has been established.