Since xylanase genes have a crucial role in degradation of xylan, a main part of plant hemicellulose, the molecular biology of such genes has become a fundamental area of research. Xylanases, secreted by such a large number of microorganisms (fungi, bacteria, actinomycetes) play a key role in biomass degradation, which is important for use in biofuel production, the management of agricultural waste, as well as for many industrial applications. This chapter focuses on the genetic and biochemical pathways present for regulation, expression and evolution of xylanase genes and the molecular mechanisms for the utilization of xylan as a carbon source by microorganisms. In addition, the section discusses the genomic and metagenomic ways to search for new xylanase genes and their regulating elements using bioinformatics tools used to predict gene functions and gene clusters. This review presents a comprehensive overview of xylanase gene molecular biology that will enable further research and technological advancements in the future to take advantage of xylanase application in biotechnology and industrial processes.