| Cherry root cancer is a worldwide bacterial disease caused by Agrobacterium tumefaciens.Carcinomas form on the roots of cherry plants,which restricts the development of the cherry industry.A non-pathogenic Agrobacterium Ag8 strain isolated from grapes in the early stage of our research group has a significant effect on the prevention and control of grape root cancer.However,whether it can effectively prevent cherry root cancer has not yet been studied.Because biocontrol agents are put into the environment in large quantities,the antibiotic resistance genes they carry will be horizontally transferred to other bacteria,which will not only produce drug resistance,but also increase the risk of threats to the environment.This study preliminarily determined the colonization of Ag8 in cherry roots and its ability to prevent cherry root cancer through pot experiments,and studied and analyzed the biological control mechanism and antibiotic resistance mechanism of Ag8 from the molecular level.Obtained the following research results:1.The good colonization ability of the biocontrol bacteria is one of the main reasons to ensure the biocontrol effect.By measuring the colonization ability of the biocontrol bacteria Ag8 in the cherry roots,the research shows that the biocontrol bacteria Ag8 is in the cherry rhizosphere soil,root surface and roots.Both can be well established,and the amount of colonization: rhizosphere soil>root surface>root inside.2.By measuring the control effect of biocontrol bacteria Ag8 on cherry root cancer and effective control measures,The results showed that when the root pathogen A.tumefaciens was used as the control,the control effect of root Ag8 on cherry root cancer was 42.87%,and the control effect of Ag8 on concrete was 40%.When the soil pathogen A.tumefaciens was used as the control,the biocontrol effect of Ag8 in dipping roots was 52.27%,and the control effect of Ag8 in irrigation roots was 20.45%.This indicated that the biocontrol bacteria Ag8 had a certain control effect on cherry root cancer,and the control effect was best when the root was dipped in Ag8.3.When the biocontrol bacteria Ag8 is put into the environment,its Kanna resistance gene may be transferred horizontally to other bacteria,causing biological environmental pollution.Therefore,this study used transposon Tn5 random insertion mutations and specific conditions to screen and construct Ag8 mutants with reduced Kanna resistance,and used TAIL-PCR to obtain the gene sequence of the insertion.The principle of two homologous recombination was used to construct the Ag8 biocontrol-related gene duf3088 and the resistance-related genes bgls,abm,exo A deletion mutants Δduf3088,Δbgls,Δabm,Δexo A and the complementing strain Δduf3088-comp,Δbgls-comp,Δabm-comp,Δexo A-comp,andΔbglsΔabm double deficiency strains were constructed on the basis of Δbgls strain.4.Through the duf3088,bgls,abm,exo A gene growth curve,motility,siderophore,biological control ability and antibiotic resistance test.The results showed that the deletion of duf3088 gene affects the prevention effect of Ag8 strain on root cancer,but does not affect the growth,motility and the ability of producing siderophore of Ag8.The bgls,abm,and exo A genes are all related to Kanna resistance,but the exo A gene has a more significant regulatory effect.In addition,the loss of exo A gene reduces the mobility and biocontrol ability of Ag8.The deletion of bgls and abm genes affects the growth and biocontrol ability of Ag8,but does not affect the mobility and the ability to produce siderophore,Compared with the Δbgls and Δabm strains,the biocontrol ability of Δbgls Δabm strain decreased more significantly. |
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