Bacteria of the genus Streptomyces are one of the most important producers of biologically active natural products. Recent robust genomic sequencing of Streptomyces strains has shown enormous genetic potential for new natural products. However, many biosynthetic gene clusters are silent. Therefore, efficient and stable genome modification methods are needed to induce their production or to manipulate them for the production of new compounds or biotechnologically improved strains. We have recently developed a simple and efficient markerless genome modification system for these bacteria based on the positive selection of double crossovers using the blue pigment indigoidine bpsA gene. This chapter is an attempt to provide methodological details of this strategy for stable markerless genomic engineering (deletions/insertions) to improve their biotechnological properties and to produce biologically active compounds.
Keywords: Antibiotic gene clusters; Gene disruption; Homologous recombination; Indigoidine; Markerless genome editing; Streptomyces.