caa a22 4500 605502137 CHVBK 20210128100604.0 cr unu---uuuuu 210128e20150501xx s 000 0 eng 10.1007/s00253-014-6251-0 doi (NATIONALLICENCE)springer-10.1007/s00253-014-6251-0 Bacilysin overproduction in Bacillus amyloliquefaciens FZB42 markerless derivative strains FZBREP and FZBSPA enhances antibacterial activity [Elektronische Daten] [Liming Wu, Huijun Wu, Lina Chen, Ling Lin, Rainer Borriss, Xuewen Gao] Bacillus amyloliquefaciens strains FZBREP and FZBSPA were derived from the wild-type FZB42 by replacement of the native bacilysin operon promoter with constitutive promoters P repB and P spac from plasmids pMK3 and pLOSS, respectively. These strains contained two antibiotic resistance genes, and markerless strains were constructed by deleting the chloramphenicol resistance cassette and promoter region bordered by two lox sites (lox71 and lox66) using Cre recombinase expressed from the temperature-sensitive vector pLOSS-cre. The vector-encoded spectinomycin resistance gene was removed by high temperature (50°C) treatment. RT-PCR and qRT-PCR results indicated that P repB and especially P spac significantly increased expression of the bac operon, and FZBREP and FZBSPA strains produced up to 170.4 and 315.6% more bacilysin than wild type, respectively. Bacilysin overproduction was accompanied by enhancement of the antagonistic activities against Staphylococcus aureus (an indicator of bacilysin) and Clavibacter michiganense subsp. sepedonicum (the causative agent of potato ring rot). Both the size and degree of ring rot-associated necrotic tubers were decreased compared with the wild-type strain, which confirmed the protective effects and biocontrol potential of these genetically engineered strains. Springer-Verlag Berlin Heidelberg, 2014 Bacillus amyloliquefaciens nationallicence Bacilysin nationallicence Promoter replacement nationallicence Markerless nationallicence Antibiotic overproduction nationallicence Antibacterial nationallicence Wu Liming Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, Nanjing, China aut Wu Huijun Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, Nanjing, China aut Chen Lina Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, Nanjing, China aut Lin Ling Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, Nanjing, China aut Borriss Rainer ABiTEP GmbH, Berlin, Germany aut Gao Xuewen Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, Nanjing, China aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/10(2015-05-01), 4255-4263 0175-7598 99:10<4255 2015 99 253 https://doi.org/10.1007/s00253-014-6251-0 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-014-6251-0 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Wu Liming Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 700 1- Wu Huijun Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 700 1- Chen Lina Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 700 1- Lin Ling Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 700 1- Borriss Rainer ABiTEP GmbH, Berlin, Germany aut NATIONALLICENCE 700 1- Gao Xuewen Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/10(2015-05-01), 4255-4263 0175-7598 99:10<4255 2015 99 253