The present study combined the fluoride ion-selective electrode （ISE） method with classical PCR to increase the detection efficiency of Burkholderia glumae. The four antisera of rabbit anti whole cell of B. glumae h...The present study combined the fluoride ion-selective electrode （ISE） method with classical PCR to increase the detection efficiency of Burkholderia glumae. The four antisera of rabbit anti whole cell of B. glumae had been successfully obtained by polyclonal antibody technique, which were ASBgl4, ASBg32, ASBg26 and ASBg04, respectively. The titers of four antisera reached 1：32 in ODD test and 1：5 120 in agglutination reaction. All antisera were identified to be qualified antibodies through combined method. The specificity of ASBgl4 and ASBg32 was very high, but that of ASBg26 and ASBg04 was not very satisfying. The results showed that all tested strains of B. glumae produced 500 bp specific fragments by direct PCR and immunocapture PCR detection, while others showed nega- tive PCR result. Comparing the sensitivity of these two detection methods, direct PCR could detect about 1 x 105 cfu/mL of bacterial suspension while immunocap- ture PCR could detect as few as 1× 103 cfu/mL, 102 times higher than direct PCR in sensitivity. In the detecting experiments of artificially inoculated seeds, at least two seeds were required in direct PCR, while only one seed was enough for immunocapture PCR detection.展开更多
文摘The present study combined the fluoride ion-selective electrode （ISE） method with classical PCR to increase the detection efficiency of Burkholderia glumae. The four antisera of rabbit anti whole cell of B. glumae had been successfully obtained by polyclonal antibody technique, which were ASBgl4, ASBg32, ASBg26 and ASBg04, respectively. The titers of four antisera reached 1：32 in ODD test and 1：5 120 in agglutination reaction. All antisera were identified to be qualified antibodies through combined method. The specificity of ASBgl4 and ASBg32 was very high, but that of ASBg26 and ASBg04 was not very satisfying. The results showed that all tested strains of B. glumae produced 500 bp specific fragments by direct PCR and immunocapture PCR detection, while others showed nega- tive PCR result. Comparing the sensitivity of these two detection methods, direct PCR could detect about 1 x 105 cfu/mL of bacterial suspension while immunocap- ture PCR could detect as few as 1× 103 cfu/mL, 102 times higher than direct PCR in sensitivity. In the detecting experiments of artificially inoculated seeds, at least two seeds were required in direct PCR, while only one seed was enough for immunocapture PCR detection.
