inSēquio: A Programmable 3D CAD Application for Designing DNA Nanostructures

Abstract DNA nanotechnology is evolving rapidly, paralleling the historic trajectory of the 1970s electronics industry. However, current DNA nanostructure (DN) design software limits users to either manual design with minimal automation or a constrained range of automated designs. inSēquio Design Studio, developed by Parabon ® NanoLabs, bridges this gap as a programmable 3D computer-aided design (CAD) application, integrating a domain-specific graphical editor with a Python API for versatile DN design. Developed in C++ for Windows ® and Macintosh ® systems, inSēquio features a user-friendly GUI with extensive CAD tools, capable of managing complex designs and offloading computational tasks to the cloud. It supports various DNA design formats, PDB molecule integration, residue modifications, and includes preloaded designs and thorough documentation. With its combination of features, inSēquio enables a code-centric design (CCD) approach, enhancing DN construction with improved precision, scalability, and efficiency. This approach is elucidated through a streptavidin barrel cage designed via Python notebook and a spheroid origami case study. Marking a significant advance in DN design automation, inSēquio, the first fully programmable 3D CAD tool for DN design, enables both manual and programmatic 3D editing. This fusion of features establishes inSēquio as a transformative tool, poised to significantly enhance designer productivity and expand the scope of possible designs. Extended Abstract Advances in DNA nanotechnology have positioned the field at a juncture reminiscent of the pivotal growth phase of the electronics industry in the 1970s. The evolution of software for designing DNA nanostructures (DNs) is following a similar historical trajectory and dozens of software packages have been developed for creating them. Existing software options, however, require users to choose between manual design with minimal automation support or selecting from a limited set of designs, typically wireframe, that can be generated from a high-level structural description. Here, we introduce the inSēquio Design Studio, a programmable 3D computer-aided design (CAD) application that effectively bridges this gap. By integrating a domain-specific, freeform graphical editor with a Python application programming interface (API), inSēquio provides a comprehensive and extensible platform for designing complex nucleic acid (NA) nanostructures. The inSēquio desktop application, developed in C++, runs on Windows ® and Macintosh ® operating systems. Its graphical user interface (GUI) features multiple synchronized view panels and a diverse set of CAD and NA-specific editing tools. Its optimized graphics pipeline enables editing of designs with >2M nucleotides, and it includes an integrated service infrastructure for offloading heavy computations to cloud servers. The software also supports import and export of various DNA design file formats, integration of arbitrary PDB molecules, and specification of residue modifications. Additionally, it includes preloaded sample designs, scripts, and comprehensive documentation. Parabon has used evolving versions of inSēquio for over a decade to design a variety of proprietary DNs and have now transitioned it into a commercially available product. This paper summarizes inSēquio’s features, discusses its strengths and limitations, and outlines planned enhancements. Although freeform 3D design is well supported in inSēquio, the integration of its CAD environment with its API facilitates a code-centric design (CCD) approach for DN construction that offers notable productivity advantages over traditional methods, including enhanced precision, scalability, and efficiency. Here we describe CCD, outline its benefits and demonstrate its use through a well-documented Python notebook, included with the product, which generates a sample design within the inSēquio application. A spheroid origami created using CCD is also presented. As the first commercial fully programmable 3D CAD application specifically created for DN design, the release of inSēquio represents a milestone in the field of DN design automation. It introduces a new dimension to the discipline by enabling both manual and programmatic 3D editing, thereby facilitating an innovative CCD approach. The availability of extensive documentation and technical support enables designers to efficiently adopt and utilize these capabilities. This combination of features establishes inSēquio as a noteworthy addition to the tools available for DN design, with the potential to significantly increase designer productivity and broaden the scope of designs that can be developed by practitioners of all skill levels. Windows and Mac versions of the inSēquio desktop application are available for download at https://parabon.com/insequio . Graphical Abstract An illustration of the inSēquio Design Studio desktop application interoperating with a Python Jupyter notebook and molecular dynamics (MD) simulation tools to support an iterative code-centric design (CCD) process. The design cycle includes (a) programmatic and/or manual creation of objects in the inSēquio editors; (b) visual inspection and manipulation of objects via user interface; (c) in silico evaluation of designs via MD simulation using native or external tools; repeating a-c as necessary; and (d) procurement of strands and synthesis of DNA nanostructures (DNs).