The advancement and accessibility of next-generation sequencing (NGS) technologies have rapidly transformed microbiological research by providing the ability to analyze and profile microbial communities via metagenomics analyses. These sequencing-based applications have relied on the availability of fully assembled reference genomes for bioinformatics analyses, particularly for variant calling in diagnostic and clinical microbiology. However, despite the availability of existing genome sequences in public databases, the quality, completeness, authenticity, accuracy, and traceability of genomic data are inadequate; the lack of standards for genome quality exacerbates these underlying problems. To address this, ATCC has implemented a robust NGS and genome assembly workflow to advance authentication of bacterial strains in the ATCC collection. Our workflow is accompanied by rigorous quality control methods and criteria to ensure that the data proceeding to the next step are the highest quality. Only data that pass all quality control criteria are published to the ATCC Genome Portal, an online database of reference-grade bacterial genomes.