Study On Efficiently Degrading Functional Genes Of Klebsiella Jilinsis To Chlorimuron-ethyl

Chlorimuron-ethyl is a ultra efficient sulfonylurea herbicide,which is widely used for the control of broad-leaved weeds in soybean fields.For its long-lasting effect and phytotoxicity to the sensitve crops,degrading chlorimuron-ethyl becomes very important.It is reported that the main degradation pathway of chlorimuron-ethyl is biodegradation and great concerns have been raised about the isolation of degrading microbes and degradation mechanisms.Klebsiella jilinsis 2N3 is a species of gram-negative bacteria that can degrade chlorimuron-ethyl and grow with chlorimuron-ethyl as the sole nitrogen source.Under appropriate condition,the degradation rate reach up to 92.5%.Furthermore,strain 2N3 can also degrade other sulfonylurea herbicides such as metsulfuron-methyl,tribenuron-methyl,rimsulfuron,ethametsulfuron and nicosulfuron.To clone the functional genes involve in the degradation of chlorimuron-ethyl by Klebsiella jilinsis 2N3,the whole genome was sequenced using the third generation high-throughput sequencing technology,and the RNA-Seq was performed using the cells of Klebsiella jilinsis 2N3 incubated with and without chlorimuron-ethyl.Analysis of genomic data showed that the genomic size of strain 2N3 was 5.33 Mb with a GC content of 42.69%.A total of 2711 coding genes were predicted and further annotated by the GO,COG,NR,and KEGG databases.Two hydrolases expressed by Kj-mhpC-2096 and Kj-mhpC-2289 were predicted to be involved in the degradation of chlorimuron-ethyl by Klebsiella jilinsis 2N3.Analysis of transcriptome data showed 30 differential expression genes in all 3 repetitions and Kj-eitD-2267?Kj-kdpD-226 and Kj-dxs-398 were predicted to be involved in the degradation of chlorimuron-ethyl.Kj-mhpC-2096,Kj-mhpC-2289,Kj-eitD-2267,Kj-kdpD-226 and Kj-dxs-398 were knocked out.The degradation activity of chlorimuron-ethyl by ?Kj-mhpC-2096 and ?Kj-dxs-398 was weakened.The degradation speed of chlorimuron-ethyl by these two mutants was significantly slower than the WT,and degradation rates of chlorimuron-ethyl by ?Kj-mhpC-2096 and ?Kj-dxs-398 were 10% lower than the WT.These findings indicate that Kj-mhpC-2096 and Kj-dxs-398 are involved in the degradation of chlorimuron-ethyl by Klebsiella jilinsis 2N3.