Article Title: Porcupine: Rapid and robust tagging of physical objects using nanopore-orthogonal DNA strands
Figure Lengend Snippet: Molbit design scheme. (a) Molecular bit (molbit) structure. The molbit sequence is attached to a spacer sequence via Golden Gate assembly to achieve a minimum length for sequencing and provide an additional encoding channel. Since the sequencing adapter is attached to both ends, the strand can be sensed from either direction. (b) The letters “UW” depicted visually in nanopore raw data (as opposed to encoded in the sequence contents). From top to bottom, the shown sequence was simulated using Scrappie and sequenced on the ONT Min-ION, demonstrating the viability of using simulations for designing intentional, arbitrary raw signal shapes. (c) Evolutionary model workflow. Each round of evolution begins with a set of sequences, their simulated squiggles, and pairwise Dynamic Time Warping (DTW) distances. The sequence order is randomized, and sequences are mutated one at a time, verifying DTW improvement (minimum and mean) after each attempt. (d) Dynamic time warping (DTW) scores before (left) and after (right) 31 iterations of the evolutionary model. After initialization, the minimum DTW similarity was 2.9 (mean 4.2 +/- 0.4), and after evolution the minimum was 4.2 (mean 5.8 +/- 0.8).
Article Snippet: To assemble the molbits, 600 ng of the desired annealed barcodes (at equimolar concentrations) and 600 ng of the spacer were ligated together using NEB’s Golden Gate Assembly Kit.