Construction of a Mycobacterium smegmatis Promoter Library for Therapeutic and Environmental Applications

Mycobacterium smegmatis is a nonpathogenic
species of soil-dwelling mycobacteria that shows promise as a synthetic
biology chassis with both clinical and environmental applications.
The development of a nonmodel chassis requires a library of regulatory
genetic elements that cover a range of expression levels. Currently,
most studied M. smegmatis promoters
are characterized using single-channel reporter cassettes, which are
vulnerable to extrinsic noise introduced by different culturing conditions,
initial cell metabolic states, and reporter genes of choice. For constructing
predictable and reliable circuits in M. smegmatis, this study systematically identified and analyzed 18 M. smegmatis promoters by using a dual-channel reporter
system across different environments. Here, we show a well-characterized
promoter library and a standardizable reporter plasmid construct that
will allow future investigators to easily assess additional promoter
elements, promoting future use of M. smegmatis as an effective and field-deployable chassis.