Bacterial mutant strains were generated as described by Baba with minor modifications [50 (link)]. Two fragments flanking the target gene were amplified from the chromosomal DNA of Xoc BLS256 using Pfu polymerase (TransGen Biotech, Beijing, China) and the primers described in S2 Table . The PCR products were digested, subcloned into the suicide vector pKMS1 [51 (link)], and introduced into bacteria by electroporation (Bio-Rad Laboratories Inc., Hercules, CA, USA) with kanamycin selection. A single transformant with kanamycin resistance was selected, cultured for 8 h in NB, and inoculated as 10-fold dilutions to NA with 15% sucrose to obtain sucrose-insensitive clones. For site-directed mutagenesis, plasmids were modified with the Fast Mutagenesis System (Transgen Biotech, Beijing, China) to obtain clones containing point mutations (Xonc3711*, 3982*, xopC2*; S1 Table ).
To obtain the Xonc3711 overexpression mutant (Xonc3711OE), the full-length corresponding gene was amplified, and the fragment was cloned into pHM1 with the lac promoter. The recombinant plasmid was transferred into WT by electroporation, and transformants were screened on NA plates supplemented with spectinomycin.
To obtain the Xonc3711 overexpression mutant (Xonc3711OE), the full-length corresponding gene was amplified, and the fragment was cloned into pHM1 with the lac promoter. The recombinant plasmid was transferred into WT by electroporation, and transformants were screened on NA plates supplemented with spectinomycin.