摘要
为了确定小麦转基因成分PCR和实时荧光PCR方法的定性检测低限,将转基因小麦B73与非转基因小麦(检测外源基因)、小麦与大米(检测内源基因)分别进行质量分数配比后提取DNA用于测定相对检测低限,再将100%转基因小麦B73的DNA进行浓度稀释用于测定绝对检测低限,并应用已知的NOS、bar、ubiquitin,ui-dA(GUS)外源基因和Wx012、GAG56 D内源基因的引物和探针对模板DNA分别进行PCR与实时荧光PCR扩增。结果表明,最终确定PCR方法检测小麦转基因成分的相对检测低限为0.1%(质量分数),绝对检测低限为0.5 ng/μL;实时荧光PCR方法的相对检测低限为0.1%(质量分数),绝对检测低限为0.01 ng/μL。所确定的检测低限可满足国家对转基因产品的最低标识要求。
This study is on the detection limits of PCR and realtime PCR for genetically modified components in wheat. B73 was the transgenic wheat line which contained ubiquitin promotor, NOS terminate, bar and uidA（GUS） reported gene, Wx012 and GAG56D endogenesis gene. Genomic DNA from transgenic wheat and non tansgenic wheat, wheat and rice （wt/wt） were extracted for the relative detection. B73 genamic DNA was tenfold serially diluted to define the absolute detection limit. After amplifing the endogenesis gene and extrinsic gene by PCR and realtime PCR, the absolute detection limit is 0.1% （wt/wt）, and the relative detection limit is 0.5 ng/uL for PCR; The absolute detection limit is 0.1%（wt/wt）, and the relative detection limit is 0.01 ng/uL for realtime PCR. The method can satisfy the expected detection limit for all the country.
出处
《麦类作物学报》
CAS
CSCD
北大核心
2009年第2期199-205,共7页
Journal of Triticeae Crops
基金
国家质检总局科研项目(2004IK084)
科技部2002年度社会公益研究专项重点项目子课题(202DIA50034-03)
国家标准委下达的国家标准制定项目(20071660-T-449)
