【摘 要】
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Activation of cryptic gene clusters in Streptomycetes requires a diversity of well-characterized regulatory elements.For characterizing the regulatory eleme
【机 构】
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ChineseAcademyofSciencesKeyLaboratoryofPathogenicMicrobiologyandImmunology,InstituteofMicrobiology,C
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Activation of cryptic gene clusters in Streptomycetes requires a diversity of well-characterized regulatory elements.For characterizing the regulatory elements, the quantitative method with single cell resolution is still lacked in such filamentous bacterium.Here, we developed a method for measuring gene expression in protoplasts of streptomycetes based on flow cytometry.The protoplasts released from the mycelium can be regarded as single cells, and the dead cells can be distinguished from the viable one by propidium iodide (PI) staining.About 200 native or synthetic promoters as well as more than 180 ribosome binding sites (RBSs) are characterized by the new method, forming a versatile toolbox for manipulating and activating the cryptic gene cluster.Furthermore, an insulator sequence (RiboJ) was employed to eliminate the interference between the promoter and RBS sequence, and improving the modularity of these regulatory elements.We believed that both the quantitative method and the two types of modular regulatory elements can facilitate the activation of the cryptic gene clusters in Streptomycetes.
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