二维电泳和iTRAQ的实验比较_潘三强.pdfVIP

Sep. 　2008　　24( 5) 　538～542 神 经 解 剖 学 杂 志 ( Ch inese Journa l of Neuroana tom y) 二维电泳和 i TRAQ的实验比较 潘三强 3 　宿宝贵 　吕来清 (暨南大学医学院　人体解剖学教研室 , 广州 510632 ) 摘　要　对蛋白组学中的二维电泳和 iTRAQ两种技术进行比较研究 ,为神经科学的蛋白组学研究提供选择参考。取生后 1～3 d C57B l/6小鼠背根神经节进行培养 ,对照组加 DMSO,实验组加 JNJ460。分别用二维电泳和 iTRAQ两种技术检测背根神经节细 胞蛋白的变化 ,用 4700 TOF /TOF串联飞行质谱仪检测变化蛋白的肽片段质量 ,用 GPS软件检索 Mascot蛋白质数据库 ,鉴定出匹 配的蛋白。利用二维电泳技术观察到 4种表达上调的蛋白 , 2种是结构蛋白 , 2种是信号转导蛋白。利用 iTRAQ技术观察到 22 种表达上调的蛋白 ,其中 10种是结构蛋白 , 3种是信号蛋白 , 2种是代谢蛋白 , 2种是降解蛋白化 , 2种是细胞基质蛋白及 3种其 它蛋白。利用二维电泳技术观察到发生表达变化的蛋白都位于胞浆内 ,而利用 iTRAQ技术观察到的表达变化的蛋白有胞浆蛋 白外 ,还有线粒体蛋白、膜蛋白和核蛋白。同时 ,利用二维电泳技术观察到的蛋白变化都在 2倍以上 ,而利用 iTRAQ技术计算出 的蛋白变化在 1. 3至 1. 6倍之间。另 , iTRAQ技术在鉴定大分子和小分子蛋白方面也有优势。结果表明 iTRAQ技术比二维电泳 技术能发现更多数量和更多种类的蛋白 ,但在比较胞浆蛋白的变化及蛋白量的变化方面二维电泳技术有一定的长处 ,对 iTRAQ 的实验结果有互补作用。 关键词　　二维电泳　iTRAQ　蛋白组学　背根神经节 A com para tive study between the two d im en siona l gel electrophoresis and iTRAQ m ethod Pan Sanqiang, Su B aogu i, LüL aiqing (Department of Anatomy, Medical College of J inan University, Guangzhou 510632) Abstract　The p resent study p rovided a reference for the neurop roteom ics study through comparison between the two dimensional gel elec2 trophoresis (22DE) and isobaric tags for relative and absolute quantification ( iTRAQ) method. The dorsal root ganglia (DRG) were cul2 tured from 1 to 3 d postnatal C57B l/6 m ice. The DRGs in control group were incubated with DMSO, and those in the experimental group were incubated with JNJ460. The pep tide masses of changed p roteins were analyzed by 4700 matrix2 assisted laser desorp tion / ionization coup led to a dual time2of2flight ( TOF /TOF) mass spectrometry, and the mass spectrometric data were used to identify the p roteins through the general p roteom ics standard ( GPS) search and Mascot database. Four p roteins were found to be up2regulated by 22DE. Two of them were structural p roteins, and the others were signaling p roteins. 22 p roteins were detected to be up2regulated from iTRAQ. These p roteins included 10 structural p roteins, 3 signalin