杨属转基因植物PCR检测内标准基因的建立

Establishment of a universal endogenous reference Gene for PCR detection of genetically modified ingredient in Populus taxa

为了建立适用于杨属植物转基因成分PCR检测的内对照系统,利用目前genebank中已公布的杨属植物的叶绿体DNArbcL基因的保守区域,设计了两对PCR引物:rbcL-1引物和rbcL-2引物,分别成功地对五大派10种杨属植物进行了rbcL基因片段的扩增,第一次建立了适用于杨属植物转基因成分PCR检测的内标准基因对照系统。其中rbcL-1引物适用的退火范围很广,在52.0～68.0℃之间均能得到特异性很强的扩增产物,这有利于其他检测项目(如35S启动子、NOS终止子、标记基因及目标基因)的引物随意搭配进行多重PCR检测,从而提高检测效率、缩短检测周期。

In order to establish a Universal endogenous reference Gene for PCR Detection of Genetically Modified Ingredient in Populns taxa, the research designed primers rbcl-1 and rbcl-2 from conservative region of rbcl gene by inquired the published sequences of Populns chloroplasts in NCBI. Fragments of rbel gene in ten species Populus classified as five sections of the genus was amplified successfully using the two primer-pairs. The endogenous reference gene control system applicabled to the transgenic Populus PCR detected was established at the first time. At the same time, the annealing temperature of primer rbcl-1 was very wide： between 52.0℃ and 68.0℃ it worked well, which will be helpfully paired up other detections stochastic, as well as improvement of detection accuracy, and reduction of detection period.