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Use of ti plasmid DNA probes for determining tumorigenicity of agrobacterium strains
T J Burr1, J L Norelli, B H Katz
1Department of Plant Pathology, New York State Agricultural Experiment Station, Cornell University, Geneva, New York 14456.
Abstract:
Probes consisting of T-DNA genes from the Ti plasmid of Agrobacterium tumefaciens were used for determining tumorigenicity of strains. Two P-labeled probes hybridized with 28 of 28 tumorigenic strains of the pathogen but not with 20 of 22 nontumorigenic strains. One probe, pTHE17, consists of all but the far left portion of the T-DNA of strain C58. Probe SmaI7 consists of SmaI fragment 7 of pTiC58, including onc genes 1, 4, and 6a and most of 2. Another probe, pAL4044, consisting of the vir region of strain Ach-5, hybridized with several nontumorigenic as well as tumorigenic strains. Colony hybridizations were done with 28 tumorigenic and 22 nontumorigenic Agrobacterium strains. About 10 CFU of the different tumorigenic strains were detectable with this method. Southern analyses confirmed the presence or absence of Ti plasmids in strains for which tumorigenicity was questioned. Colony hybridization with the T-DNA probes provides a rapid and sensitive means for determining the tumorigenic nature of Agrobacterium strains.
Insights
Researchers developed a rapid method using T-DNA probes to identify tumorigenic Agrobacterium strains. This technique accurately distinguishes between disease-causing and non-disease-causing strains, aiding in plant disease diagnostics.
Area of Science:
- Microbiology
- Plant Pathology
- Molecular Biology
Background:
- Agrobacterium tumefaciens causes plant tumors via its Ti plasmid.
- Accurate identification of tumorigenic strains is crucial for disease management.
Purpose of the Study:
- To develop and validate a rapid method for determining the tumorigenicity of Agrobacterium strains.
- To assess the efficacy of T-DNA probes for strain differentiation.
Main Methods:
- Utilized P-labeled T-DNA probes derived from the Ti plasmid of Agrobacterium tumefaciens.
- Performed colony hybridization assays on 28 tumorigenic and 22 nontumorigenic Agrobacterium strains.
- Confirmed results using Southern blot analyses for Ti plasmid presence.
Main Results:
- Two specific T-DNA probes hybridized with all 28 tumorigenic strains but only 2 of 22 nontumorigenic strains.
- The method detected as few as 10 colony-forming units (CFU) of tumorigenic strains.
- A probe targeting the vir region showed less specificity, hybridizing with both strain types.
Conclusions:
- Colony hybridization with T-DNA probes offers a sensitive and rapid diagnostic tool for Agrobacterium tumorigenicity.
- This method can reliably differentiate between pathogenic and non-pathogenic strains.
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