Publication highlights

Researchers at the Crick have developed SubtiToolKit (STK), a high-efficiency Golden Gate (GG) toolkit - a method for molecular cloning - for Bacillus subtilis and other Gram-positive bacteria, addressing a key gap in synthetic biology tools. It includes a GeoBox for Geobacillus spp. engineering, which demonstrates versatility for other bacteria. The GG method allows a rapid modular construction of complex genetic circuits by exploiting Type IIS restriction enzymes and standardised overhangs (single strands of DNA extended from a double-stranded DNA fragment produced by restriction enzymes). The STK is designed with high-efficiency overhangs that enable precise assembly of transcriptional units (TU), operons, and constructs for genomic integration, reducing time, cost, and experimental complexity. It includes libraries of promoters, ribosome-binding site (RBSs), terminators, protein tags, fluorescent proteins, antibiotic cassettes, and tools to deal with toxic constructs during the assembly process. The STK unlocks the cutting-edge methods for bioengineering in Gram-positive bacteria.