As more and more systems biology approaches are used to investigate the different types of biological macromolecules, increasing numbers of whole genomic studies are now available for a large array of organisms. Whether it is genomics, transcriptomics, proteomics, interactomics or metabolomics, the full complement of genomic information on all different levels can be juxtaposed between different organisms to reveal similarities or differences, and even to provide consensus models. At the intersection of comparative genomics and systems biology lies great possibility for discovery, analysis and prediction. This paper explores this nexus and the relationship from four general levels: DNA, RNA, protein and extragenomic. For each level, we provide an overview of the methods, discuss the potential challenges and survey the current research. Finally, we suggest some organizing principles and make proposals for new areas that will be important for future research.