Single-base mismatch recognition using partially double-stranded probes having various lengths.

We have focused on the nucleation-synchronized DNA strand exchange reaction (ns-SER) as a format of a single-base mismatch detection. We employed a partially double-stranded (PDS) DNA probe having a single-stranded (ss) portion. We could rapidly resolve a single-base mismatch by the ns-SER rates of the PDS probe with target DNAs. Here, we reported the influence of probe length and operating conditions, such as temperature and buffer conditions, on a single-base mismatch recognition using the PDS probes. We could reliably recognize a single-base mismatch even with a 61mer long probe. However, the ns-SER rates were decreased with increasing probe length. We considered that a rate-limiting step of the ns-SER might be sifted from nucleation to branch migration processes. By an addition of cationic comb-type copolymers (PLL-g-Dex) in the buffer, the reaction rate was considerably increased 1-2 orders without disordering the resolution power.