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试验用PCR方法从玉米中扩增内源基因zSSIIb片段,并将其克隆到pMD18-T载体上,获得中间载体pMD-zSSIIb;根据Bt11和MON810玉米转化体特异性序列,分别设计带酶切位点的引物,扩增出Bt11和MON810转化体特异性产物;用相应的酶对pMD-zSSIIb和两种PCR产物进行酶切,分别将Bt11和MON810产物克隆到pMD-zSSIIb上,获得阳性标准分子pMD-ZB和pMD-ZM,并对其进行特异性测试。结果表明,获得的阳性标准分子可以作为转基因产品检测时的阳性对照。
The zSSIIb fragment of endogenous gene was amplified from maize by PCR and cloned into pMD18-T vector to obtain the intermediate vector pMD-zSSIIb. Based on the specific sequences of Bt11 and MON810 maize transformants, Point primers were used to amplify specific products of Bt11 and MON810 transformants; pMD-zSSIIb and two PCR products were digested with the corresponding enzymes, and the Bt11 and MON810 products were respectively cloned into pMD-zSSIIb to obtain positive standard molecules pMD-ZB and pMD-ZM, and tested for specificity. The results showed that the obtained positive standard molecule can be used as a positive control when the transgenic product is detected.