16S rDNA BASED IDENTIFICATION OF BUTACHLOR DEGRADING SOIL BACTERIAL ISOLATES FROM RICE FIELD IN UTTARAKHAND TERAI REGION
Neha Trivedi, A. K. Verma, Ashutosh Dubey
Organic pesticide accumulation in the field causes ammonia and nitrite poisning which create water and soil pollution. It becomes essential to do efforts regading removal of such waste. Micro-organisms play important role in removal of such compounds by evolving their metabolisms. In this study culturable bacterial biodiversity of butachlor degrading four bacterial isolates indigenous to soil herbicide was assessed. These soil bacterial isolates were characterized through a variety of phenotypic, morphologic, biochemical and molecular properties. Molecular identification of these bacterial isolates was done through partial 16s rDNA based gene sequencing method. Bacterial 16S ribosomal RNA (rRNA) genes contain nine “hyper-variable regions” (V1–V9) that demonstrate considerable sequence diversity among different bacteria. PCR Amplification of 16S rDNA of variable regionV4-V5 for four isolates BC1, BC2, BC3 and BC4 were carried out by using universal primers and products were sequenced commercially. These gene sequences were compared with other gene sequences in the GenBank database. A phylogenetic tree was constructed. It was found that isolates belonged to different species of genus Bacillus, sharing 92-99% 16S rDNA identity, isolates also had close similarities with Penibacillus and Geobacillus with 97%, 98% 16S rDNA. Gene for partial 16s rRNA were deposited to GenBank and accession numbers were obtained. Based on this study it can be concluded that, soil having continuous exposure to butachlor is populated with diverse bacterial groups. Partial 16s rDNA sequencing can be used to identify the microflora responsible for degradation of hazardous pesticides.
Keywords: Biodiversity, Characterization; Butachlor, PCR amplification of 16S rDNA.