摘要
玉米组织蛋白质的提取及其浓度测定是进行玉米蛋白质组学研究的关键步骤。本研究以玉米叶片和花丝为试材,从总蛋白提取数量和质量方面比较分析了磷酸缓冲液法、Tris法、改进Tris法、TCA-acetone(三氯醋酸-丙酮沉淀)法、Plant Total Protein Extraction Kit试剂盒法及其改良法等6种玉米总蛋白质提取方法。同时,又比较分析了Bradford法、2-D Quant Kit试剂盒法和紫外分光光度法(UV法)等3种方法测定蛋白质浓度的准确性。结果表明:(1)总蛋白提取数量按大小顺序为:试剂盒法>试剂盒改良法>改进Tris法>Tris法>TCA-acetone法>磷酸缓冲液法;(2)总蛋白提取质量依质量优劣为:试剂盒法>试剂盒改良法>TCA-acetone法>改进Tris法>Tris法>磷酸缓冲液法;(3)Bradford法与2-D Quant Kit法测定的蛋白质浓度均是准确的,且没有显著差异;但是UV法测定的蛋白质浓度欠准确。经综合考虑,改进Tris法用于SDS-PAGE分析,试剂盒改良法用于2-DE分析,Bradford法用于测定蛋白质浓度是经济有效的方法。这对玉米蛋白质组学的研究具有参考价值。
The extraction and concentration determination of total protein in maize organs is a key step for proteomic research.In this study.Using maize leaves and silks as materials,6 methods for total protein extraction,i.e.Phosphate Buffer（PB）,Tris,Improved Tris,TCA-acetone,Plant Total Protein Extraction Kit and its improved one were compared in respective extraction quantity and quality;simultaneously,3 methods of Bradford,2-D Quant Kit and UV were compared in accuracy of protein concentration determination.The results showed：1.the amount of total protein extracted by the above methods was Plant Total Protein Extraction KitImproved Plant Total Protein Extraction Kit〉Improved Tris〉Tris〉TCA-acetone〉 PB;2.The quality of total protein extracted was Plant Total Protein Extraction KitImproved Plant Total Protein Extraction Kit〉TCA-acetone〉 Improved Tris〉Tris〉PB;3.The protein concentrations determined by both methods of Bradford and 2-D Quant Kit were accurate and no significant difference,however the one determined by UV method was inaccurate.Taking one consideration with another,using Improved Tris for SDS-PAGE and Improved Plant Total Protein Extraction Kit for 2-DE analyses,and Bradford for concentration measurement would be efficient.It should be a good reference for proteomic research in maize.
出处
《种子》
CSCD
北大核心
2010年第12期18-23,共6页
Seed
基金
国家重点基础研究发展计划(973计划)项目(2006CB708206):玉米奥帕克-16(o16)和果穗性状的表达分析及o16基因的克隆
贵州省农业科学院专项资金项目(黔农科院院专项[2009]032号):玉米奥帕克-16突变的蛋白质表达分析
贵州省科学技术基金(黔科合J字[2010]2082号):玉米奥帕克-16突变的蛋白质表达分析
关键词
蛋白质提取
蛋白质浓度测定
玉米
protein extraction
protein concentration determination
maize
