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作 者:钱昱霏[1] 刘建国[1] 白国辉[1,2] 管晓燕[1] 韩琪[1] 白朋元[1]
机构地区:[1]遵义医学院口腔学院,贵州遵义563099 [2]遵义医学院医学与生物学研究中心,贵州遵义563099
出 处:《遵义医学院学报》2013年第2期116-120,共5页Journal of Zunyi Medical University
基 金:国家自然科学基金项目(NO:30160086;81260164);贵州省优秀科技人才省长专项基金项目(NO:黔省专合字[2006]44);贵州省科技攻关项目(NO:黔科合SY字[2012]3086);贵州省科技厅联合基金项目(NO:黔科合J字LKZ[2011]41)
摘 要:目的比较两种不同配方的蛋白提取液提取转基因番茄蛋白的效果。方法用A、B两种植物蛋白提取液提取相同防龋用转基因番茄蛋白,运用western blot、BCA法和间接ELISA法对提取出的转基因番茄外源目的蛋白进行检测。结果两种提取液提取的蛋白样品均在相对分子质量约为60 KDa处出现了低分子量蛋白条带,但植物蛋白提取液B所提取蛋白样品条带更清晰。蛋白提取液A、B分别提取的相同转基因番茄总蛋白为8.375mg/mL和12.838 mg/mL,而其外源目的蛋白的含量分别是0.335μg/mL和1.16μg/mL,比值为2.25倍。经重复实验验证B液提取的目的蛋白在总蛋白含量的百分比较A液所提取的高,具有统计学意义(P<0.05)。结论植物蛋白提取液B提取转基因番茄中外源目的蛋白较提取液A更为高效并且省时。Objective To compare two kinds of extraction method of transgenic tomato expressing target protein by optimizing the reagent formula. Methods Two kinds of plant protein extracting solution A and B were applied to extract genetically modified tomato protein for caries prevention. Western blot analysis, BCA assay and indirect ELISA method were performed to test exogenous protein level of transgenic tomato. Results The protein samples extracted from two extracting solution were all appeared The low density protein bands with relative molecular weight of 60 KD were shown in the protein sam- ples extracted from two extracting solution. The extracted protein sample bands of plant protein extracting solution B were thicker and more distinct than those in solution A. The total protein contents of transgenic tomato extracted from solution A and B were 8. 375 mg/mL and 12. 838 mg/mL, while exogenous target protein levels were 0. 335μg/mL and 1.16μg/mL, respectively, with the ratio of 2. 25 times. The percentage of purpose protein in total protein extracted from solution B through repeated experiments was more than that from solution A ( P 〈 0.05 ). Conclusion The extraction method using plant protein extracting solution B is an efficient and timesaving mthod of transgenic tomato expressing target protein extraction.
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