Burkholderia sp. have been isolated from the rhizosphere soils of different plants, viz., maize, sugarcane, red gram, rice, bhendi (Okra), brinjal, cotton, chili, cowpea etc. The isolates were checked for nitrogen fixation, ability to solubilize insoluble phosphate and antagonistic activity against plant pathogenic fungi Rhizoctonia solani and Macrophomina phaseolina by using different growth media like BMGM, HAP, PDA+NA respectively. The isolates were also checked for nitrogenase activity by using gas chromatography. B1 and R1 isolates showed high nitrogenase activity (63.45 and 62.35 n moles of ethylene/hr./ml respectively). Total genomic DNA was isolated and PCR with 16S rRNA gene specific primers yields amplicons of 1.5 kb size in 4 selected isolates of Burkholderia sp. The PCR was carried out with specific primers of nif H gene primer for selected isolates. It showed that selected four isolates were found to have nif H gene with 400 bp. Sequence analysis revealed homology of 16S rRNA amplicons of B1 as Burkholderia thailandensis and R1 as Burkholderia vietnamiensis. The present study revealed that some of the Burkholderia sp. helps in plant growth promoting activities like nitrogen fixation, phosphate solubilization and antagonism to enhance the yield of crop plants that can be exploited as bioinoculant in agriculture.