On March 16, 2018, the U.S. Food and Drug Administration (FDA) published an advance notice of proposed rulemaking (ANPRM) entitled “Tobacco Product Standard for Nicotine Level of Combusted Cigarettes.” The publication of an ANPRM solicits information and public comments from stakeholders –industry, public health advocates, etc. – regarding a particular issue. After all of the requested information has been submitted, the FDA must consider the information in fashioning a proposed rule, or it may decline to issue a proposed rule. This presentation centers on one aspect of the referenced ANPRM – establishing a nicotine level standard in combustible cigarettes. Specific potential nicotine levels mentioned in the ANPRM are: 0.3, 0.4, and 0.5 mg nicotine/g of tobacco filler. Can levels this low be achieved in the tobacco plant and maintain tobacco leaf yield and quality? Low nicotine tobacco lines containing the nic1/nic2 loci have been available for several years, but agronomic data from large field trials is scarce and nic1/nic2 will not achieve the lowest levels mentioned in the ANPRM. Molecular biology techniques have been employed to elucidate information for many of the major nicotine biosynthesis pathway genes as well as transporters, transcription factors and non-coding RNAs. By providing an overview of these various genetic aspects and their feasibility, insight into some of the challenges facing the tobacco industry will be presented. Challenges are to not only comply with the potential nicotine level standard, but maintain both flavor characteristics acceptable to adult tobacco consumers as well as crop yields and tobacco leaf quality tobacco producers require.