MicroRNAs (miRNAs) belonging to the same family have similar sequences and are difficult to identify. Herein, we report the reverse transcription-hairpin-probe-polymerase chain reaction (RT-Hpro-PCR) technique, which utilises a reverse transcription (RT) primer containing a 5′-end deoxyribonucleic acid (DNA) tag, to detect miRNAs with similar sequences. This strategy follows a two-step RT-PCR method using 6–7-mer RT-primers with a ~ 10-mer tag sequence at the 5′-end and a probe with a hairpin structure (Hpro), including two C-bulges, attached. The findings demonstrate that the specificity of RT could be increased by shortening the complementary part of the RT primer containing a different base, wherein the PCR could successfully progress with the use of 5′-end DNA tag because of an increase in the length of the hybridised tagged primer. This study shows the potential of RT-Hpro-PCR to precisely detect miRNAs with similar sequences, which could help explore the roles of miRNAs in several biological processes.

属于同一家族的微小RNA（miRNA）具有相似的序列并且难以识别。在此，我们报告了逆转录-发夹-探针-聚合酶链反应 (RT-Hpro-PCR) 技术，该技术利用含有 5' 端脱氧核糖核酸 (DNA) 标签的逆转录 (RT) 引物来检测具有相似的序列。该策略遵循两步 RT-PCR 方法，使用 6–7 聚体 RT 引物，在 5' 端具有 ~ 10 聚体标签序列，以及具有发夹结构 (Hpro) 的探针，包括两个 C 凸起， 随附的。研究结果表明，RT 的特异性可以通过缩短含有不同碱基的 RT 引物的互补部分来增加，其中 PCR 可以通过使用 5' 端 DNA 标签成功进行，因为增加了长度杂交标记引物。