转基因玉米pUC57-BT11质粒标准分子的构建与适用性研究

摘要

构建了含有转化体特异性片段( BT11-93bp)和玉米的内标准基因片段(zSSⅡb-151 bp)的质粒分子pUC57-BT11,经酶切和测序验证后,对其特异性进行检查.采用测序和荧光定量PCR法对该质粒分子进行定值,结果为1.01±0.01(k=2).通过质粒分子和基因组DNA产生的内源和外源基因比较,发现两者标准曲线斜率无显著差异,线性相关系数均≥0.99.利用pUC57-BT11质粒分子对已知含量的转基因基体标准物质进行转基因含量测定,结果发现采用pUC57-BT11质粒分子对标准物质的测定结果与其标准值一致,表明pUC57-BT11质粒分子可作为转基因玉米BT11转化体特异性的定性和定量检测用标准物质.%To construct a plasmid pUC57-BTll including event specific product and endogenous gene zSSIIb product After restriction enzyme digestion and sequencing, the specific of the plasmid pUC57-BTll is detected. The property value of pBTl 1 is 1.01 ± 0. 01 ( k = 2 ) determined by sequencing and real time PCR. By comparison standard curve generated by plasmid with genomic DNA, no significant difference between the slopes of the standard curve is found. All correlated coefficient R2 are ≥0. 99. Certified GM matrix materials are detected by quantitative PCR when pBTll plasmid DNA is used as quantification standards. The results determined by quantitative PCR with pBTl 1 are identical with the certified values. This indicated that pBTl 1 plasmid can be used as a standard for CM maize BT11 quantitative and qualitatively detection.