This chapter discusses recent genetic and metabolic engineering advances to produce liquid biofuel from renewable biomass. It provides a comprehensive overview of the techniques employed, including metabolic pathway engineeringMetabolic pathway engineering, strain improvement and synthetic biologySynthetic biology approaches. In this chapter, discussion on metabolic pathway that involves gene expression, gene know-out and also introduction to a novel pathway to enhance biofuel productionBiofuel production. In addition, the potential of microorganisms’Microorganisms strain improvement as well as synthetic biologySynthetic biology to design new strain catalyst to produce liquid biofuel such as bioethanolBioethanol, biobutanolBiobutanol and biodieselBiodiesel from biomass are evaluatedBiomass. The chapter highlights several investigations, demonstrating the potential of these technologies to produce sustainable and renewable drop-in biofuelsDrop-in biofuels. The future of genetic and metabolic engineering for liquid biofuel productionBiofuel production appears promising to be an alternative to partially replace energy from fossil fuel. This chapter also discusses the challenge that potentially arises from the development of the liquid biofuel from genetically modified microorganismsMicroorganisms. Overall, this chapter serves as a comprehensive insight into production of liquid biofuel, challenges, and future directions through genetic and metabolic engineering.