Poster: Biology & Disease Mgmt: Biological Control
Developing a quantitative polymerase chain reaction protocol to quantitate root colonization by Bacillus amyloliquefaciens and Bacillus firmus
H. MENDIS (1), L. De La Fuente (1), P. Schwientek (2), R. Salamzade (2), V. Thomas (2), J. Kloepper (3) (1) Auburn University, U.S.A.; (2) Bayer CropScience LP, U.S.A.; (3) Auburn University, U.S.A.
Bacillus amyloliquefaciens QST713 and B. firmus I-1582 are 2 key bacterial strains which are active ingredients in commercially-available biological products that colonize plant roots promoting plant growth and offering protection against pathogens. This study was carried out to develop a qPCR protocol to understand the dynamics of root colonization by these strains. Primers and TaqMan probes were designed based on genome comparisons of the 2 strains with other publicly available bacterial genomes. The specificity of the primers was tested in vitro and in growth chambers focusing on corn rhizosphere. An optimized qPCR protocol was developed to quantify the number of bacteria in batch cultures and in root samples using standard curves. Corn seeds were treated with 103, 105 and 107 spores of the 2 strains separately and planted in sterile sand. Whole root samples were taken at 2 and 3 weeks after germination and bacterial populations were assessed. QST713 and I-1582 primers did not amplify DNA extracted from untreated corn grown soil confirming that the primers are specific. The results also show that as low as 3.5x103 bacteria cells associated with 1g of corn roots can be detected using the current qPCR protocol. Furthermore, the number of bacteria cells associated with corn roots was quantified up to 3 weeks after germination. Our results show that the developed qPCR protocol can be used to understant dynamics of root colonization by plant growth promoting bacteria.