AbstractMost of Earth’s bacteria have yet to be cultivated. The metabolic and functional potentials of these uncultivated microorganisms thus remain mysterious, and the metagenome-assembled genome (MAG) approach is the most robust method for uncovering these potentials. However, MAGs discovered by conventional metagenomic assembly and binning are usually highly fragmented genomes with heterogeneous sequence contamination. In this study, we combined Illumina and Nanopore data to develop a new workflow to reconstruct 233 MAGs—six novel bacterial orders, 20 families, 66 genera, and 154 species—from Lake Shunet, a secluded meromictic lake in Siberia. With our workflow, the average N50 of reconstructed MAGs greatly increased 10–40-fold compared to when the conventional Illumina assembly and binning method were used. More importantly, six complete MAGs were recovered from our datasets. The recovery of 154 novel species MAGs from a rarely explored lake greatly expands the current bacterial genome encyclopedia.