Molecular Identification of Soil Bacteria by 16srDNA Sequence

Abstract

In this current study, 16S rDNA (genotypic) identification technique is focused on identification of conventionally unidentifiable isolates those are unevaluated in isolated by employing molecular techniques and Bioinformatics in uploading and retrieving isolate gene sequences which are rapid, reliable and accurate in differentiation of various soils isolates. This study is an automaton of 16Sr DNA gene sequence that allows a queue comparison analysis of published sequences deposited in the microbial genome database was used. Polymerase chain reaction (PCR) amplification of 16SrDNA gene using the consensus bacterial primer and separation of the resulting polymer chain reaction amplicon  by cloning, temperature gradient electrophoresis are major ecological techniques that are used in the description of soil bacteria. The isolated gene was cloned using PTZ57r or T cloning Vector amplified using 16SF and 16SR primer transformed in DH5? Cells resulting PCs 16s Plasmid hybrid. The primer 16S F₂ obtained from M13 forward primer was used and aligned using BLAST and submitted to EMBL+GENEBANK+DDBJ+ PDB. 99% similarity was observed and later it was analyzed with the existing sequence in ribosomal database project II.  RDP classifier was used for confirmation with 100 % similarity. The bacteria were identified as Burkholderia cenocepacia  when the  sequence was submitted and retrieved via the World Wide Web and new sequence compared with those held in the database  using the basic local alignment tool (BLAST). A segment of 734 out of 736 nucleotide of 16S rDNA gene of Burholderia Cenocepacia is the region of choice for primer construction because of proximity that provides a successful discrimination in strains of Burholderia Cenocepacia in soil. 16S rDNA gene account to 99%  similarity score in molecular typing and identification of bacteria which concerns deposition of sequences into established microbial genomic database

Key Words: Burkholderi; Bacterial transformation; Characterization; DNA based techniques