Identification of Genes Involved in Antifungal Activity of Burkholderia seminalis Against Rhizoctonia solani Using Tn5 Transposon Mutation Method
Abstract
:1. Introduction
2. Results and Discussion
2.1. Construction of Tn5 Mutant Library
2.2. Location of Tn5 Inserted Sites
2.3. Functional Prediction of Antagonism-Related Genes
2.4. Verification by Constructing Deletion Mutants and Complements
2.5. Growth of Antagonism-Related Tn5 Transposon Mutants
2.6. Biofilm Formation
2.7. Motility
2.8. Tolerance to Hydrogen Peroxide (H2O2)
3. Materials and Methods
3.1. Bacterial Strains and Growth Conditions
3.2. Transposon Insertion of Tn5
3.3. Evaluation of Antagonistic Activity of Tn5 Mutants
3.4. Identifying the Location of Transposon Insertions
3.5. Generation of Deletion Mutants and Complemented Strains
3.6. Growth Measurement
3.7. Biofilm Formation Ability
3.8. Motility
3.9. Tolerance to H2O2
3.10. Statistical Analyses
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Conflicts of Interest
Abbreviations
BLAST | Basic local alignment search tool |
IR | Index of resistance |
PCR | Polymerase chain reaction |
PDA | Potato dextrose agar (medium) |
ShB | Rice sheath blight |
WT | Wild type |
References
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Mutants | Locus_Tag | Enzyme Site | Sequence Length (bp) | Function Prediction |
---|---|---|---|---|
Tn5-30 | BsemR456_1725 | SalI | 861 | glycosyl transferase |
Tn5-45 | BsemR456_1417 | PaeI | 203 | multispecies: histone H1 |
Tn5-63 | BsemR456_2057 | SalI | 479 | nonribosomal peptide synthetase |
Tn5-145 | BsemR456_1423 | SalI | 55 | 50S ribosomal protein L11 methyltransferase |
Tn5-146 | BsemR456_6327 | SalI | 150 | Plasmid, membrane integrity-associated transporter subunit PqiC |
Tn5-158 | BsemR456_6210 | SalI | 452 | Plasmid, hypothetical protein |
Tn5-216 | BsemR456_1157 | SalI | 765 | tRNA uridine-5-carboxymethylaminomethyl (34) synthesis enzyme MnmG. |
Tn5-225 | BsemR456_2679 | PaeI | 33 | YeiH family putative sulfate export transporter |
Tn5-273 | BsemR456_1556 | SalI | 281 | catalase/peroxidase HPI |
Tn5-331 | BsemR456_315 | SalI | 1173 | sulfate adenylyltransferase subunit CysD |
Tn5-355 | BsemR456_6210 | SalI | 452 | Plasmid, hypothetical protein |
Bacterial Strains | Incubation Time (h) | |||
---|---|---|---|---|
6 | 12 | 18 | 24 | |
Wild type | 0.50 ± 0.03 ab | 0.90 ± 0.00 c | 1.12 ± 0.00 bc | 1.31 ± 0.00 ab |
Tn5-30 | 0.34 ± 0.00 d | 0.90 ± 0.08 c | 1.14 ± 0.02 b | 1.25 ± 0.04 b |
Tn5-45 | 0.53 ± 0.01 ab | 0.93 ± 0.05 bc | 1.09 ± 0.02 c | 1.27 ± 0.02 b |
Tn5-63 | 0.40 ± 0.02 c | 0.81 ± 0.03 d | 1.07 ± 0.02 c | 1.28 ± 0.01 b |
Tn5-145 | 0.56 ± 0.06 a | 1.06 ± 0.02 a | 1.19 ± 0.03 a | 1.36 ± 0.02 a |
Tn5-146 | 0.47 ± 0.08 b | 0.99 ± 0.04 ab | 1.12 ± 0.03 bc | 1.31 ± 0.00 ab |
Tn5-158 | 0.49 ± 0.03 b | 0.99 ± 0.03 b | 1.12 ± 0.04 bc | 1.28 ± 0.08 b |
Tn5-216 | 0.30 ± 0.01 d | 0.73 ± 0.03 e | 0.90 ± 0.03 e | 1.10 ± 0.06 c |
Tn5-225 | 0.47 ± 0.04 b | 0.98 ± 0.06 b | 1.05 ± 0.01 d | 1.04 ± 0.00 c |
Tn5-273 | 0.46 ± 0.03 bc | 0.95 ± 0.04 bc | 1.10 ± 0.02 c | 1.32 ± 0.03 ab |
Tn5-331 | 0.44 ± 0.01 bc | 0.93 ± 0.01 bc | 1.11 ± 0.03 bc | 1.28 ± 0.05 b |
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Zhang, M.; Wang, X.; Ahmed, T.; Liu, M.; Wu, Z.; Luo, J.; Tian, Y.; Jiang, H.; Wang, Y.; Sun, G.; et al. Identification of Genes Involved in Antifungal Activity of Burkholderia seminalis Against Rhizoctonia solani Using Tn5 Transposon Mutation Method. Pathogens 2020, 9, 797. https://doi.org/10.3390/pathogens9100797
Zhang M, Wang X, Ahmed T, Liu M, Wu Z, Luo J, Tian Y, Jiang H, Wang Y, Sun G, et al. Identification of Genes Involved in Antifungal Activity of Burkholderia seminalis Against Rhizoctonia solani Using Tn5 Transposon Mutation Method. Pathogens. 2020; 9(10):797. https://doi.org/10.3390/pathogens9100797
Chicago/Turabian StyleZhang, Muchen, Xiaoxuan Wang, Temoor Ahmed, Mengju Liu, Zhifeng Wu, Jinyan Luo, Ye Tian, Hubiao Jiang, Yanli Wang, Guochang Sun, and et al. 2020. "Identification of Genes Involved in Antifungal Activity of Burkholderia seminalis Against Rhizoctonia solani Using Tn5 Transposon Mutation Method" Pathogens 9, no. 10: 797. https://doi.org/10.3390/pathogens9100797