Micro RNAs (miRNAs) are involved in a variety of biological functions and are attracting attention as diagnostic and prognostic markers for various diseases. Highly sensitive RNA detection methods are required to determine miRNA expression levels and intracellular localization. In this study, we designed new double-stranded peptide nucleic acid (PNA)/DNA probes consisting of a fluorophore–PNA–quencher (fPq) and a quencher–DNA (qD) for miR-221 detection. We optimized the fPq structure, PNA–DNA hybrid length, and hybrid position. The resultant fPq-2/qD-6b probe was a 6-bp hybrid probe with a 10-base fPq and a 6-base qD. The signal-to-background ratios of the probes showed that fPq-2/qD-6b had a higher target sensitivity than fPq (PNA beacon)-type and fP/qD-type probes. The results of the detection limit and target specificity indicate that the fPq/qD probe is promising for RNA detection in both cells and cell extracts as well as for miRNA diagnosis.

微小 RNA (miRNA) 涉及多种生物学功能，并作为各种疾病的诊断和预后标志物而受到关注。需要高度灵敏的 RNA 检测方法来确定 miRNA 表达水平和细胞内定位。在这项研究中，我们设计了由荧光团-PNA-猝灭剂 (fPq) 和猝灭剂-DNA (qD) 组成的新型双链肽核酸 (PNA)/DNA 探针，用于检测 miR-221。我们优化了 fPq 结构、PNA-DNA 杂交长度和杂交位置。得到的 fPq-2/qD-6b 探针是一个 6-bp 的混合探针，具有 10 个碱基的 fPq 和一个 6 个碱基的 qD。探针的信背景比表明，fPq-2/qD-6b 的目标灵敏度高于 fPq（PNA 信标）型和 fP/qD 型探针。