【摘要】 根据转基因油菜中所转入的外源基因,选择CaMV35S启动子、FMV35S,PAT基因和目的基因mCP4-EP-SPS,BAR,MS8,RF3以及内源PEP基因设计特异性引物,采用多重PCR法对待测样品进行扩增,通过缺口平移法合成DIG-dUTP标记杂交探针,并制备基因芯片,在对PCR反应和扩增产物与芯片杂交条件进行优化的同时,比较了芯片检测的特异性和重复性,并对检测的灵敏度进行测试,结果表明,该方法具有较好的特异性和重复性,检测灵敏度可达0.1%,由于采用了多重PCR技术一次可同时检测多个基因,提高了检测的准确性和效率.更多还原

【Abstract】 According to the plasmid map of genetically modified rapeseed,some specificity primers were designed by selecting exogenous genes,e.g.CaMV35S promoter,FMV35S promoter,PAT gene,mCP4-EPSPS gene,BAR gene,MS8 gene,RF3 gene and endogenous gene(PEP) as target genes.The hybridized probe labeled with DIG-dUTP was synthesized using the nick translation method.Multiplex PCR was used to amplify the target sequence in maize sample DNA chip,and then the conditions of hybridization for DNA chips with PCR product was optimized.The results showed that the DNA chip for identifying genetically modified rapeseed was highly specific and repeatable and its sensitivity was 0.1%,meanwhile the detection efficiency was highly improved by using multiplex PCR.更多还原