Complementation and introducing other genes into S. vietnamensis

For complementation or introducing other genes into S. vietnamensis, the target genes were amplified and then assembled individually, using the ClonExpress MultiS One Step Cloning Kit, into pSETAT-KasOp* between the AflII and SpeI sites where the open reading frame (ORF) was immediately downstram the promotor KasOp*. The resulting plasmids were then individually introduced into the Δgra-orf32 mutant or other strains with the same method as described in the in-frame deletion section. Screening of the desired colonies followed the standard procedure. The E. coli FAS ACPS gene and the PPTase gene sfp of surfactin biosynthesis of Bacillus subtilis were codon-optimized and synthesized by Azenta China (Supplementary Figures S1,2). The PPTase genes jadM and med-orf24 of jadomycin and medermycin biosynthetic pathways were also synthesized since these biological materials are not available in our lab. The synthesized genes were cloned into pSETAT-KasOp* between the AflII and SpeI sites as same as other genes.