Short hairpin RNAs of designed sequences can be extracellularly produced by the marine bacterium Rhodovulum sulfidophilum

Previously, we proposed a new method for production of RNA aptamers using the marine bacterium Rhodovulum sulfidophilum. A streptavidin RNA aptamer (an RNA which binds to streptavidin) was extracellularly produced by this bacterium containing engineered plasmid. The aptamer had full biological function. As a next step we attempted to produce another functional RNA, short hairpin RNAs (shRNAs) using this bacterial system. We have designed two types of shRNAs targeted to the luciferase gene. Here we report that shRNAs are successfully produced extracellularly by this system. Even if the shRNA has a long stem-loop structure which is thought to interfere with transcription in bacterial cells, the yield of the shRNA is almost the same as that of the streptavidin RNA aptamer. During the course of these experiments, we also found a new type of RNA processing for the double-stranded region of the shRNA.