Figure 5. Recognition and control of DNA complexes in real time using FPGA.

a, A representative ternary complex event under FPGA control. The FPGA samples the ionic current every 5.3 μs and computes a windowed mean amplitude (solid dark line) based on the previous 1.5 ms of signal. Every 0.4 ms, it tests whether or not the mean is within 24 ± 2.8 pA. i, If the mean remains within this range for four consecutive tests, the FSM logic diagnoses the blockade as a KF binding event. The total delay for diagnosis of a KF binding event is 2.7 ms (1.5 ms for the windowed mean to enter the 24 ± 2.8 pA range, plus 1.2 ms for three consecutive subsequent tests). ii, Upon diagnosis of a KF binding event, the FPGA continues to monitor the windowed mean. If the mean remains in the 24 ± 2.8 pA range for 20 ms, the FSM logic diagnoses the blockade as resulting from a ternary complex. The 20 ms cutoff was used because 60% of events are longer than 20 ms in the presence of the correct dNTP, but only 2% of events are longer than 20 ms in the 24 ± 2.8 pA range in the absence of the correct dNTP. iii, Upon diagnosis that a ternary complex is in the pore, the FPGA reverses the voltage to −50 mV for 5 ms, ejecting the complex from the pore. The 180 mV capture voltage is then restored. For each event in which KF binding is not diagnosed, or in which KF binding is diagnosed but the mean leaves the 24 ± 2.8 pA range before 20 ms, the voltage is kept at 180 mV. b, Dwell time probability histograms for 24 ± 2.8 pA events with FPGA control (527 total events, in red) and without FPGA control (155 total events, in blue).