Evaluation of Genetic Switches and Reporters as Synthetic Biology Tools for Clostridium acetobutylicum
Abstract
This report presents an evaluation of reporter proteins and inducible gene expression elements in Clostridium (C.) acetobutylicum, a bacterium of interest to the Army. Reporter proteins evaluated were LacZ (beta-galactosidase), PhiLOV, Evoglow C-Bs2, SNAP-Tag, FAST, and NanoLuc luciferase. Inducible systems evaluated included six riboswitches and seven chemically inducible promoter systems. Most of these genetic tools had not been previously tested in C. acetobutylicum. The theophylline-inducible riboswitch and the NanoLuc luminescent reporter protein proved useful, convenient, and modular. The FAST and SNAP-Tag fluorescent reporters also proved worthwhile for flow cytometry. A subset of these genetic tools was deployed as part of a study and evaluation of four different Gram-positive origins of replication in C. acetobutylicum: pBP1, pCB102, pCD6, and pIM13, with respect to their utility for elevated protein expression. The pIM13 origin yielded the highest level of protein expression, which was about 20-fold greater than that conferred by pBP1, the origin with the next-highest copy number. An outlook perspective is provided, based on this and related work, of genetic tools that could be tested or retested in C. acetobutylicum, with lessons from this work as a guide.
Document Details
- Document Type
- Technical Report
- Publication Date
- Dec 01, 2019
- Accession Number
- AD1088634
Entities
People
- Alexander V. Tobias