一种用于核酸高灵敏检测的液滴式数字聚合酶链式反应芯片  

Novel Microfluidic Droplet Digital PCR Chip for High Sensitive Detection of Nucleic Acid

作　　者：袁浩钧 郜晚蕾 景奉香[1] 刘松生 周洪波[1] 贾春平[1] 金庆辉[1] 赵建龙[1] 

机构地区：[1]传感技术国家重点实验室中国科学院上海微系统与信息技术研究所,上海200050 [2]中国科学院大学,北京100049

出　　处：《高等学校化学学报》2017年第7期1140-1147,共8页Chemical Journal of Chinese Universities

基　　金：国家自然科学基金(批准号:81472751;61271162;61571428;61571429);国家"九七三"计划项目(批准号:2012CB933303);上海市浦江人才计划项目(批准号:15PJ1409800)资助~~

摘　　要：为了实现对核酸的高灵敏度检测,构建了一种新型的液滴式数字聚合酶链式反应(dd PCR)芯片.该芯片由产生液滴的聚二甲基硅氧烷(PDMS)模块和储存液滴的玻璃腔室构成.实验结果表明,该芯片可以在25 min内产生2×106个直径为20μm的微液滴(体积4.187 p L).利用该芯片定量检测了表皮生长因子受体(EGFR)基因第19号外显子,在DNA浓度为106~101copies/μL范围内呈现良好的线性关系(R2=0.9998);在浓度为106copies/μL的19号外显子野生型DNA中检测105~100copies/μL的突变型DNA,其检测敏感度可达到0.0001%.该方法在同一芯片上实现了液滴产生、核酸扩增和荧光信号读取的功能,在核酸绝对定量及痕量突变基因的检测中具有潜在应用前景.Sensitive and rapid quantification of nucleic acid is significant for clinical diagnosis and medication guide of tumor. In this study, we developed a novel droplet digital PCR（ddPCR） device for sensitive nucleic acid detection. This chip contains a PDMS chip and glass-made reservoir, which can overcome the problems of droplet evaporation and instability during PCR. The double droplet generation channels in this chip can produce 2×106 droplets（4. 187 pL each） in 25 min. For detecting EGFR exon 19 wild-type DNA using this ddPCR chip, it demonstrated a good linear correlation with the DNA concentration from 10 to 106 copies/μL （R2= 0. 9998）. In addition, we show sensitive detection of EGFR exon 19 mutate DNA in a 106-fold excess of wild-type bac PCR process nucleic acids kground. In conclusion, this ddPCR chip integrates the functions of droplet generation, on-chip and fluorescence signals readout, which will provide a potential application in the absolute quantification and rare mutate gene detection.

关 键 词：数字聚合酶链式反应 微流控芯片 微液滴 高灵敏度核酸检测 

分 类 号：O655.9[理学—分析化学]