Development of a semi-nested PCR based method for sensitive detection of tumorigenic Agrobacterium in soil.
OBJECTIVE
To develop a specific, sensitive and rapid PCR-based method for detection of tumorigenic Agrobacterium in soil.


RESULTS
Three newly designed primers complementary to tms2 gene amplified DNA of only the tumor-inducing agrobacteria of 113 strains tested, resulting in 617 bp and 458 bp products in the first and second rounds of semi-nested PCR respectively. As optimized method of pre-incubation of soil suspensions on selective medium, DNA isolation and two-round semi-nested PCR enabled detection of 1-2 bacterial cells in 1 g of soil. Using this method tumour-inducing Agrobacterium was detected in 67 of 69 samples of naturally infested soil originating from the field, where plants with crown gall symptoms occurred. The pathogen was detected only in two samples of 15 tested, collected from a nursery where crown gall symptoms were not observed.


CONCLUSIONS
The semi-nested PCR-based method allowed for sensitive and rapid detection of tumorigenic agrobacteria in soil.


CONCLUSIONS
The method is proposed for testing of soil in fields intended for nursery production of fruit trees, roses or other plants susceptible to crown gall, as well as a tool for ecological studies.
Puławska
J
J
Research Institute of Pomology and Floriculture, Skierniewice, Poland. jpulaw@insad.pl
Sobiczewski
P
P
D006368:Helianthus / Q000382:microbiology; D036741:Plant Tumor-Inducing Plasmids; D016133:Polymerase Chain Reaction / Q000379:methods; D012231:Rhizobium / Q000235:genetics / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D012988:Soil Microbiology