AbstractBenthic microbial mats dominated by Cyanobacteria are important features of polar lakes. Although culture-independent studies have provided important insights into their diversity, only a handful of genomes of polar Cyanobacteria have been sequenced to date. Here, we applied a genome-resolved metagenomics approach to data obtained from Arctic, sub-Antarctic, and Antarctic microbial mats. We recovered 22 unique metagenome-assembled genomes (MAGs) of Cyanobacteria, most of which are only distantly related to genomes that have been sequenced so far. These include i) lineages that are common in polar microbial mats such as the filamentous taxaPseudanabaena, Leptolyngbya, Microcoleus/Tychonema, andPhormidium; ii) the less common taxaCrinaliumandChamaesiphon; iii) an enigmatic Chroococcales lineage only distantly related toMicrocystis; and iv) an early branching lineage in the order Gloeobacterales that is almost exclusively restricted to the cold biosphere, for which we propose the nameCandidatusSivonenia alaskensis. Our results show that genome-resolved metagenomics is a powerful tool for expanding our understanding of the diversity of Cyanobacteria, especially in understudied remote and extreme environments.Data summaryThe sequencing data generated in this study have been submitted to the European Nucleotide Archive (ENA) under the BioProject PRJEB59431. Individual accession numbers for raw reads and genomic bins are listed inTable S1andTable S3, respectively. Genomic bins can also be downloaded from doi.org/10.6084/m9.figshare.22003967. The commands used throughout this study are available ingithub.com/igorspp/polar-cyanobacteria-MAGs.Impact statementCyanobacteria are photosynthetic microorganisms that play important roles in polar lacustrine ecosystems. Many Cyanobacteria are difficult to grow in the laboratory, particularly in isolation from other organisms, which makes it challenging to sequence their genomes. As such, considerably fewer genomes of Cyanobacteria have been sequenced so far compared to other bacteria. In this study, we used a metagenomics approach to recover novel genomes of Cyanobacteria from Arctic and Antarctic microbial mats without the need to isolate the organisms. The community DNA was extracted and sequenced, and the genomes of individual populations were separated using bioinformatics tools. We recovered 22 different genomes of Cyanobacteria, many of which have not been sequenced before. We describe in more detail an interesting lineage of ancestral Cyanobacteria in the order Gloeobacterales, for which we propose the nameCandidatusSivonenia alaskensis. Our study shows that genome-resolved metagenomics is a valuable approach for obtaining novel genomes of Cyanobacteria, which are needed to improve our understanding of life in the polar regions and the planet at large.