CAJ | 학술논문

为建立稳定有效检测miR-34c的实时荧光定量技术体系，采用茎环结构法设计了反转录引物，并通过梯度浓度及梯度温度PCR法对miR-34c的最佳退火延伸温度和模板浓度进行了确定，最后通过实时荧光定量 PCR对miR-34c进行检测。试验成功建立了精确检测miR-34c的实时荧光定量技术体系，其中miR-34c最佳退火延伸温度为62．4℃及每个实时定量反应体系中 cDN A 对应的 RN A 量最适浓度为50 ng/μL ，并得到重复性高、特异性强的检测结果。该技术体系的建立不仅为检测miR-34c提供了精准便捷的方法，而且可为设计检测其他miRN A提供技术支持。
위건립은정유효검측miR-34c적실시형광정량기술체계，채용경배결구법설계료반전록인물，병통과제도농도급제도온도PCR법대miR-34c적최가퇴화연신온도화모판농도진행료학정，최후통과실시형광정량 PCR대miR-34c진행검측。시험성공건립료정학검측miR-34c적실시형광정량기술체계，기중miR-34c최가퇴화연신온도위62．4℃급매개실시정량반응체계중 cDN A 대응적 RN A 량최괄농도위50 ng/μL ，병득도중복성고、특이성강적검측결과。해기술체계적건립불부위검측miR-34c제공료정준편첩적방법，이차가위설계검측기타miRN A제공기술지지。
To set up a real-time qPCR technology to detect miR-34c effectively and stably ,this article used stem-loop structure method to design the reverse transcription primers ,and then through concentration gradient and temperature gradient PCR method ,we identified the best miR-34c extend annealing tempera-ture and template concentration .Finally ,we detected miR-34c by using real-time quantitative PCR .Final-ly ,this experiment successfully established a real-time qPCR with the best annealing temperature of 62 .4℃and the appropriate RNA concentration of 50 ng/μL which turn into cDNA through inverse tran-scription in each reaction ,and acquired the high repeatability and specificity .In conclusion ,the establish-ment of the technical system not only provide an accurate and convenient method for the detection of miR-34c ,but also offer technical supporting to design and detect other miRNA .