Identification and Characterization Auxin Producing Bacteria From the Rhizosphere of Wheat.

Growth and development of wheat is highly dependent on microbial interactions in the rhizosphere. A subset of rhizosphere bacteria are known to produce indole-3-acetic acid (auxin), a major plant hormone. These auxin producing bacteria (APB) promote both growth and disease. Here we have isolated and identified 75 APB to level of genera from the rhizosphere of wheat by partially sequencing the 16s rRNA gene. Wheat was grown in a sandy loam soil for 4 weeks, roots harvested with soil still clinging and bacteria extracted in 0.1 % sodium pyrophosphate by homogenization. The extract was centrifuged at slow speed (800g) and the supernatant serially diluted and plated onto 1/10x TSA. Over 1500 colonies were randomly picked, triple steak purified, grown in liquid TSA and archived at -80 C in ATCC freezer medium. DNA was extracted using MoBio UltraClean DNA purification kit and the DNA was stored at -80 C. The 16s rRNA gene was amplified using the 27f forward and 23s reverse primers, and the 27f primer was used to sequence an approximately 800 bp region. Sequence data was compared to 16s rRNA type sequences stored in the Ribosomal Project Database II for putative identification to the level of genera. Wheat seed was inoculated with each APB and planted in sandy and clay loam soil to test their plant growth promotion or reduction capabilities. Nineteen genera were identified with a Shannon and Simpson diversity index of 2.62 and 0.078, respectively. Bacillus (22.2%), Flavobacteria (13.3%), Pseudomonas (8.9%) were the most prevalent among all 75 APB. Brevundimonas, Aminobacter, Brevibacillus, Caulobacter, and Pseudoxanthomonas were shown for the first time to be auxin producers. We conclude that APB from wheat rhizosphere is more diverse than previously estimated.