A programmable nucleic acid fluorescence biosensor based on the BstNI endonuclease for detection of the influenza A...

Timely and accurate diagnosis of influenza virus is essential to prevent the spread of disease and to select an appropriate treatment strategy. Here, we report the development of a novel fluorescence biosensor for detection of the H1N1 virus based on the BstNI endonuclease, dually-blocked RNA strands (S), and FAM-ssDNA-Q reporter (R) strands. The S strand contains a short (5 nt) sequence for BstNI recognition and sequences on both sides of the cutting site, which are closed by two locking strands. The R strand is complementary to the intermediate sequence of the S strand, including the BstNI cutting site domain and partial adjacent sequences. Only the presence of two specific RNA fragments of the target influenza virus can fully de-block the S strands, which then hybridize with the R strands, followed by cleavage via BstNI-catalyzed hydrolysis, thereby generating the fluorescence signal. The biosensor was sensitive to the H1N1 virus at 100 fM and was highly specific to the target sequence. The proposed biosensor provides a convenient method for reliable detection of the H1N1 virus.