分子生物学手段检测转Bt基因棉花中Cry杀虫蛋白的定性和定量方法

Molecular Biological Methods for Qualitative and Quantitative Analysis of Cry Insecticidal Proteins in Transgenic Bt Cotton

中文关键词:  转Bt基因棉花  Cry杀虫蛋白  酶联免疫法  凝胶电泳  实时荧光定量PCR

英文关键词:transgenic Bt cotton  Cry insecticidal protein  ELISA  gel electrophoresis  real-time fluorescence quantitative PCR

中文摘要:

      建立了转Bt基因棉花中Cry杀虫蛋白的提取、样品前处理以及酶联免疫（ELISA）定量分析方法，并使用凝胶电泳、普通聚合酶链式反应（PCR）和实时荧光定量PCR等分子生物学手段对转基因棉花中的Bt基因进行定性和定量检测。所建立的苏云金芽孢杆菌杀虫晶体蛋白（Cry1Ab蛋白和Cry1Ac蛋白）标准曲线线性关系良好，相关系数r²均大于0.999，相对标准偏差RSD均小于2.0%。方法简单、快速、重现性和精密度好，可为农业食品行业和环境领域科研人员提供一种简便快速地从转基因棉花

英文摘要:

      An enzyme linked immunosorbent assay(ELISA) quantitative method has been established to analyze Cry proteins extracted from transgenic Bt cotton. Various molecular biological methodologies such as gel electrophoresis, ordinary polymerase chain reaction(PCR) and real-time fluorescence quantitative PCR were applied to detect Bt gene in transgenic cotton qualitatively and quantitatively. Both standard curves of Bacillus thuringiensis insecticidal crystal protein Cry1Ab and Cry1Ac proteins in the ELISA analysis were of good linearity with correlation coefficients(r²) of > 0.999, and the relative standard deviations(RSD) for the analyses were less than 2%. The newly established method is easy to operate, rapid, reproducible and precise, and it can be used in the fields of agro-food industry and environmental science to analyze Bt toxin proteins in genetically modified crops.