Isolation, Characterization And Identification Of Nitrate Reductase-Producing Bacteria

Research Article
Marwa Eltarahony, Sahar Zaki, Zainab hassan Khairalla and Desouky Abd-El-Haleem
DOI: 
xxx-xxxxx-xxxx
Subject: 
science
KeyWords: 
Nitrate reductase, napA, narG, Achromobactersp., characterization
Abstract: 

Due to the environmental importance of nitrate-reductase enzyme (NR) in removing nitrate overload, this study aimed to screen, isolate and characterize NR producing bacteria. To achieve this target, various soil and water samples were collected from different Egyptian ecosystems. Isolation of NR producing bacteria was performed on BTB medium. About 43 different isolates were selected. In order to reduce the large number and eliminate similar isolates 16S rDNA-RFLP approach employed toreduced them from 43 to 25.The hyper-active bacterial isolate has been identified by physiological, microscopically and biochemical tests and16S rDNA sequencing as Achromobacter and named Achromobacter sp. strain MMT. It reached the maximum enzyme activity after 12 h, total removing of nitrate after 48h and complete nitrite conversion to N2 gas after 90 h of incubation. Detection, sequencing and phylogenetic analysis of narG and napA genes in strain MMT were carried out using nested PCR with specific primers and the presence of both genes was confirmed. Amplified nar G and nap A PCR products were TA cloned and analyzed by sequencing. In addtion, determination of NR activity in PAGE zymograme was perfomed confirming the ability of strain MMT to reduce nitrate under aerobic and anaerobic conditions, respectively.