n-day-old fresh roots of seedlings were collected straight in the plates and washed briefly in sterile water in preparation for scanning electron microscope (SEM) imaging. Roots have been cut into 5-mm lengths and fixed inside a three glutaraldehyde buffered with 0.1 M Bcl-xL Inhibitor MedChemExpress phosphate buffer (pH 7.0) for 24 h at four C. Root samples have been then completely rinsed in 0.1 M phosphate buffer (pH 7.0) and dehydrated at 25 C working with a graded ethanol series (25, 50, 75, 85, and 100 ethanol). Final, the samples had been dried with a essential point dryer, sputter-coated with platinum, and viewed in SEM (Jeol, Tokyo, Japan). Single strain B2 was also observed applying SEM. Briefly, following incubation in LB for 48 h at 30 C, strain B2 was collected by centrifugation. Immediately after washing 3 times with phosphate buffer, strain B2 was fixed with 3 glutaraldehyde in phosphate buffer at four C for 24 h. Just after washing 3 occasions with phosphate buffer,Identification of B. amyloliquefaciens BThe traditional physiological and biochemical traits of strain B2 had been identified determined by Bergey’s Manual of Systematic Bacteriology. Strain B2 was additional identified by way of the evaluation of its 16S rDNA and gyrB gene sequences. Briefly, the genomic DNA with the strain B2 was extracted utilizing the bacterial DNA extraction kit (Omega, Germany) and stored at 0 C. The 16S rDNA was amplified with all the bacterial universal primers 27F (five -AGAGTTTGATCCTGGCTCAG-3 ) and 1492R (five -GGTTACCTTGTTACGACTT-3 ) (Eden et al., 1991), and the gyrB gene was amplified with the certain primers UP1 (five GAAGTCATCATGACCGTTCTGCAYGCNGGNGGNAARTTY GA-3 ) and UP2r (5 -AGCAGGGTACGGATGTGCGAGCCRT CNACRTCNGCRTCNGTCAT-3 ) (Yamamoto and Harayama, 1995). The 20- PCR mixture contained 2 dNTP (2 mM), 2 MgCl2 (25 mM), 1.0 of every single primer (10 mM), 2.0 PCR buffer (10, 1.0 template DNA, 0.2 Taq DNA polymerase (five U), and ten.8 double-distilled (dd) H2 O. The CB1 Inhibitor drug thermocycling procedure involved an initial denaturation at 95 C for 3 min, followed by 35 cycles at 95 C for 1 min, 50 C for 45 s, 72 C for 2 min, plus a final extension at 72 C for ten min. The PCR items were then purified and sequenced by Majorbio Bio-pharm Technology Co., Ltd. (Shanghai, China). A sequence similarity analysis was performed utilizing the NCBI BLAST program1 , and also the phylogenetic tree was constructed by the neighbor-joining (NJ) system applying MEGA-X.http://blast.ncbi.nlm.nih.gov/Blast.cgiFrontiers in Microbiology | frontiersin.orgAugust 2021 | Volume 12 | ArticleWang et al.Co-application of Bacteria and FungusFIGURE 2 | Antagonism of B. amyloliquefaciens B2 against plant pathogen F. oxysporum f. sp. cucumerinum (FOC). (A) Antagonistic effects of strain B2 against FOC. (B) FOC grown on potato dextrose agar (PDA) plate as manage.the samples have been dehydrated applying a graded series of ethanol solutions (25, 50, 75, 85, and 100 ethanol). They were then dried, sputter-coated, and viewed with the SEM.60, 72, 84, and 96 h and freezing the samples at 0 C for later evaluation. The fungal mycelia biomass and residual phenolic acid concentrations have been detected as described above.Identification of Optimal Concentration for P. ostreatus P5 DegradationTo study the effects of unique initial concentrations of mixture of phenolic acids [p-hydroxybenzoic acid, vanillic acid, ferulic acid, p-coumaric acid, benzoic acid (1/1/1/1/1, w/w)] on degradation, 2-ml inocula containing 1.2 mg L-1 of mycelia were added to 50-ml mineral salt medium (MSM; KCl 0.5 g, K2 HPO4 1 g, KNO3 two g, Mg
