两种Taqman探针的制备及荧光检测性能比较

摘要

采用固相合成法,分别合成FAM/TAMRA和FAM/BHQ2双荧光标记的Taqman探针,得到的粗产物分别采用乙醇沉淀法、离子对反相色谱法、凝胶过滤柱脱盐法纯化.采用实时荧光PCR法,分别从感染的南瓜果实中检测到黄瓜绿斑驳花叶病毒(CGMMV),扩增曲线表明采用该实验条件制备的探针检测性能良好.在相同荧光PCR反应体系下,通过比较△Rn、CT值,证明FAM/BHQ2探针检测效果好于FAM/TAMRA探针.%By adopting solid-phase synthesis,synthesize Taqman probes with double fluorescence-labeled FAM/TAMRA and FAM/ BHQ2 respectively. The obtained crude products purified by applying methods of ethanol precipitation,reversed-phase ion-pair chro-matography and gel-filtration desalting. By real time fluorescence PCR,the amplified curves showed that the probes work well by comparing the results of respectively detected Cucumber Green Mottle Mosaic Virus (CGMMV)from infected pumpkin fruit Under the same fluorescent PCR reaction system,by comparing the △Rn and Cτ value the result showed FAM/BHQ2 is better than the probe FAM/TAMRA.