Exploring novel RNA topologies is imperative for understanding RNA structure and pursuing its design. Our RNA-As-Graphs (RAG) approach exploits graph theory tools and uses coarse-grained tree and dual graphs to represent RNA helices and loops by vertices and edges. Only dual graphs represent pseudoknotted RNAs fully. Here we develop a dual graph enumeration algorithm to generate an expanded library of dual graph topologies for 2 to 9 vertices, and extend our dual graph partitioning algorithm to identify all possible RNA subgraphs. Our enumeration algorithm connects smaller-vertex graphs, using all possible edge combinations, to build larger-vertex graphs and retain all non-isomorphic graph topologies, thereby more than doubling the size of our prior library to a total of 110,667 dual graph topologies. We apply our dual graph partitioning algorithm, which keeps pseudoknots and junctions intact, to all existing RNA structures to identify all possible substructures up to 9 vertices. In addition, our expanded dual graph library assigns graph topologies to all RNA graphs and subgraphs, rectifying prior inconsistencies. We update our RAG-3Dual database of RNA atomic fragments with all newly identified substructures and their graph IDs, increasing its size by more than 50 times. The enlarged dual graph library and RAG-3Dual database provide a comprehensive repertoire of graph topologies and atomic *

RNA molecules are composed of modular architectural units that define their unique structural and functional properties. Characterization of these building blocks can help interpret RNA structure/function relationships. We present an RNA secondary structure motif and submotif library using dual graph representation and partitioning. Dual graphs represent RNA helices as vertices and loops as edges. Unlike tree graphs, dual graphs can represent RNA pseudoknots (intertwined base pairs). For a representative set of RNA structures, we construct dual graphs from their secondary structures, and apply our partitioning algorithm to identify non-separable subgraphs (or blocks) without breaking pseudoknots. We report 56 subgraph blocks up to nine vertices; among them, 22 are frequently occurring, 15 of which contain pseudoknots. We then catalog atomic fragments corresponding to the subgraph blocks to define a library of building blocks that can be used for RNA design, which we call RAG-3Dual, as we have done for tree graphs. As an application, we analyze the distribution of these subgraph blocks within ribosomal RNAs of various prokaryotic and eukaryotic species to identify common subgraphs and possible ancestry relationships. Other applications of dual graph partitioning and motif library can be envisioned for RNA structure analysis and design.

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