Its. Eighteen chosen strains had been assessed for siderophore CLK review production based on
Its. Eighteen chosen strains were assessed for siderophore production according to the O-CAS method [17]. Phosphate-solubilizing activity was tested on Pikovskaya CYP1 Source medium [18], NBRIP medium [19] and modified Burk’s agar medium [1], adding 0.5 of Ca3 (PO4 )two to each and every medium as insoluble P supply. In both assays, Pseudomonas fluorescens2. Supplies and Methods2.1. Soil Sampling, Bacterial Isolation, and Azotobacter Reference Strains. In total, 74 bulk soil samples (00 cm) had been collected from agricultural (53 samples) and non-agricultural web pages (21 samples) during spring 2006. Samples belonged to 38 different areas of Northwest, Pampas, and Patagonia regions of Argentina (see Supplementary Material obtainable on the net at dx.doi.org/10.1155/2013/519603). Soil aggregates (two mm) were spread onto the surface of Petri dishes containing N-free Burk’s agar medium with mannitol as C-source [1]. Just after 5 days at 28 C, slimy and glistening Azotobacter-like colonies expanding about soil particles have been selected and further purified in N-free LG with bromothymol blue agar medium [1]. Motility, pigment production, and encystment have been determined as previously described [1].The Scientific World Journal BNM233 (Banco Nacional de Microorganismos, Buenos Aires, Argentina) was made use of as a optimistic control. Auxin production was determined employing a colorimetric assay [20], with measurements soon after 1, 2, three, and 5 days of development in modified LG (LGSP) liquid medium containing 1 sucrose and 0.five soymeal peptone. At every time interval, the amount of cells (cfu mL-1 ) was determined by plate counting on LG agar. Nitrogenase activity was estimated by the acetylene reduction assay. Bacterial cultures were grown in N-free Burk’s agar medium at 28 C for 24 h and ethylene production was measured by gas chromatography [21], applying a Hewlett Packard Series II 5890 equipped with a flame ionization detector (FID) along with a stainless-steel Porapak N column (three.2 mm two m; 80/100 mesh). The injector, oven, and detector temperatures had been 110 C, 90 C, and 250 C, respectively. N2 was employed as carrier gas (four.five cm s-1 linear gas velocity). Total protein concentration of bacterial cells was determined by the Lowry technique with all the DC Protein Assay kit (BioRad, USA). Nitrogenase activity was expressed as mmol ethylene created per mg of protein in 24 h. Indole-3-acetic acid (IAA), gibberellic acid (GA3 ), and zeatin (Z) production had been determined for six chosen Azotobacter spp. strains grown in LGSP liquid medium at 28 C for eight days. Z was identified and quantified by HPLC-UV, whereas IAA and GA3 had been identified by gas chromatography-mass spectrometry with selective ion monitoring (GC-MS-SIM), as previously described [21]. two.7. Effects of Azotobacter Inoculation and IAA Pure Solutions around the Quantity of Seminal Roots and Root Hairs of Wheat Seedlings. For plant tests, seeds of wheat (Triticum aestivum cv. Baguette Premium 13, Nidera, Buenos Aires, Argentina) have been surface-disinfected (1 NaClO for three minutes) and germinated in plastic containers (15 25 4 cm) on filter paper soaked with sterile distilled water. To preserve humidity, containers had been wrapped in transparent plastic bags and placed in a growth chamber at 25 C using a 16 h light/8 h dark regime for 24 h. For inoculation, bacterial strains were grown in LGSP liquid medium at 28 C for 8 days (108 cfu mL-1 ). Fifteen pregerminated seeds have been inoculated with one hundred L of bacterial culture (107 cells) per seed and grown for eight days as described ab.
