Direct Readout of Single Nucleobase Variations in an Oligonucleotide

Direct, low‐cost, label‐free, and enzyme‐free identification of single nucleobase is a great challenge for genomic studies. Here, this study reports that wild‐type aerolysin can directly identify the difference of four types of single nucleobase (adenine, thymine, cytosine, and guanine) in a free DNA oligomer while avoiding the operations of additional DNA immobilization, adapter incorporation, and the use of the processing enzyme. The nanoconfined space of aerolysin enables DNA molecules to be limited in the narrow pore. Moreover, aerolysin exhibits an unexpected capability of detecting DNA oligomers at the femtomolar concentration. In the future, by virtue of the high sensitivity of aerolysin and its high capture ability for DNA oligomers, aerolysin will play an important role in the studies of single nucleobase variations and open up new avenues for a broad range of nucleic‐acid‐based sensing and disease diagnosis. An aerolysin nanopore directly achieves the identification of four types of single nucleobase (adenine, thymine, cytosine, and guanine) in a free DNA oligomer without the use of additional operations. The high sensitivity of aerolysin nanopores for DNA detection opens up new avenues for nucleic acid analysis and provides opportunities for a wide range of nucleic‐acid‐based sensing.