化学学报 ›› 2004, Vol. 62 ›› Issue (3): 274-278. 上一篇    下一篇

研究论文

复合式蝎形引物实时定量检测端粒酶延伸产物

黄艳萍1,2, 孔德明1, 张晓滨1, 沈含熙1, 宓怀风1   

  1. 1. 南开大学化学学院高分子功能材料吸附与分离国家重点实验室, 天津, 300071;
    2. 天津医科大学药学院, 天津, 300070
  • 投稿日期:2003-09-24 修回日期:2004-01-08 发布日期:2014-01-26
  • 通讯作者: 沈含熙,E-mail:hxshen@eyou.com E-mail:hxshen@eyou.com
  • 基金资助:
    国家自然科学基金(No.20075012)资助项目.

Real-time Quantitative Assay of Telomerase Product Using the Duplex Scorpion Primer

HUANG Yan-Ping1,2, KONG De-Ming1, ZHANG Xiao-Bin1, SHEN Han-Xi1, MI Huai-Feng1   

  1. 1. State Key Laboratory of Functional Polymer Materials for Adsorption and Separation, Chemical School, Nankai University, Tianjin 300071;
    2. College of Pharmacy, Tianjin Medical University, Tianjin 300070
  • Received:2003-09-24 Revised:2004-01-08 Published:2014-01-26

针对端粒酶延伸产物中靶基因序列的特殊性,开发了一种可产生荧光的复合式蝎形引物,该引物的5'端带有可特异性检测靶基因的探针序列,PCR阻断剂将其与引物序列连接.当复合式蝎形引物延伸,探针序列与同一分子内的靶基因杂交,荧光信号产生.运用该技术,建立了定量检测端粒酶延伸产物的实时荧光PCR方法.该法可在快速PCR循环条件下,对0.15~1.50×103 amol/μL范围内的样品进行定量检测,线性相关系数R2=0.9992.该法操作简便,无需PCR后额外的检测步骤.

关键词: 复合式蝎形引物, 端粒酶, 实时检测, 聚合酶链式反应(PCR)

To specific target sequence of telomerase product, a fluorogenic duplex scorpion primer has been designed. A probe element attached at the 5'-end of it can specifically detect target gene. A PCR blocker—carbon chain with C3 group, whose length is as the same as a base pair, is joined between the primer sequence and the probe in the duplex scorpion primer. A fluorescence signal is only produced when the probe sequence is hybridized with the target gene in the extended duplex scorpion primer. Using this technology, a novel method has been developed for quantitative assay of telomerase product by real-time PCR. Accurate quantitative assay can be achieved with sample detected within 0.15~1.50×103 amol/μL under fast cycling conditions. The linear correlation factor R2=0.9992. The method is specific, simple and without post-PCR manipulation.

Key words: duplex scorpion primer, telomerase, real-time assay, polymerase chain reaction(PCR)