General Cloning – Geobacillus
The pMTL60000 plasmids are modular shuttle vectors for use in Parageobacillus thermoglucosidasius (formerly Geobacillus glucosidasius) and other Gram-positive thermophiles. They essentially comprise four modular parts that are combined through four, unique restriction enzymes with 8bp recognition sequences: AscI, FseI, PmeI and SfbI.
Parts comprise a Gram +ve replicon that functions in the thermophilic host, a Gram-ve replicon that supports plasmid maintenance in the E. coli shuttle host, a selectable marker, and an application specific module. Several of the replicons include origins of transfer that support mobilisation of the vector by conjugation. Parts are arranged relative to one another in the same order and orientation. Each exists as at least three variants – with a current total of 20 parts available1. These are distributed as seven plasmids, which together contain all 20 modules and are available from the SBRC online shop.
- Sheng, L., Kovacs, K., Winzer, K., Zhang, Y., and Minton N. P. (2017) Development and implementation of rapid metabolic engineering tools for chemical and fuel production in Geobacillus thermoglucosidasius NCIMB 11955. Biotechnol. Biofuels 2017; 10, 5, https://doi.org/10.1186/s13068-016-0692-x
- Madika A, Spencer J, Lau MSH, Sheng L., Zhang Y., and Minton N. P. (2022) pMTL60000: A modular plasmid vector series for Parageobacillus thermoglucosidasius strain engineering. J. Microbiol Methods 202; 106600, https://doi.org/10.1016/j.mimet.2022.106600