刺苋和苋属的实时荧光PCR鉴定

摘要

基于叶绿体的PsbA基因(Genbank登录号:DQ006133.1),设计了ASP-1和ASP-2两组具有刺苋特异性的引物探针,对苋科Amaranthaceae苋属Amaranthus刺苋A.spinosus的9个不同地理种、长芒苋A.palmeri的2个不同地理种、其它5种苋属植物和苋科非苋属8种,以及苋科外植物3种共27个样品进行TaqMan实时荧光PCR鉴定.结果表明,ASP-1适合作为刺苋的实时荧光PCR鉴定引物,其最佳扩增温度62℃;ASP-2适合作为苋属的实时荧光PCR鉴定,其最佳扩增温度为60℃.首次建立了TaqMan探针实时荧光PCR快速鉴定刺苋和苋属植物的方法.在优化后的检测条件下,ASP-1的检测灵敏度可达到0.03 ng·μL-1;ASP-2的检测灵敏度则可达到0.01 ng·μL-1.上述方法作为形态鉴定的辅助方法,可准确有效的对刺苋及苋属植物进行鉴定.%Based on the conserved fragment of plant chloroplast genes (Genbank No. DQ006133.1), a primer-probe set specific for Amaranthus spinosus (ASP-1) and a primer-probe set specific for Amaranthus spp. (ASP-2) were designed. Screened by to the 27 species including 9 target species, 7 closely-related species and 11 other non-target species,the two primer-probe sets show good specificity. According to the real-time PCR tests results, the optimum amplification temperatures for ASP-1 and ASP-2 are 62℃ and 60℃ separately. Under optimal conditions ASP-1 and ASP-2 showed high efficiency in the detection of target plants with the detection limits of 0.03 ng·μL-1 for A.spinosus and 0.01 ng·μL-1 for Amaranthus spp., respectively. It is the first time to establish a real-time PCR method for rapid identification of Amaranthus spinosus and Amaranthus spp., and as an auxiliary method of morphological identification, it can be implemented accurately and effectively.