2.3.4. ACC Deaminase Activity

For determining the ACC deaminase activity, endophyte strains were grown in sterilized minimal Dworkin and Foster (DF) salt media (DF salt contains 6.0 g Na₂HPO₄, 4.0 g KH₂PO₄, 0.2 g MgSO₄·7H₂O, 2.0 g citric acid, 2.0 g glucose, and 2.0 g gluconic acid per liter with the following trace elements: 124.6 mg ZnSO₄·7H₂O, 78.22 mg CuSO₄·5H₂O, 11.19 mg MnSO₄·H₂O, 10 mg MoO₃, 10 mg H₃BO₃, 1 mg FeSO₄·7H₂O; pH 7.2) amended with 3 mM ACC instead of (NH₄)₂SO₄ as a sole nitrogen source [44,45]. The inoculated plates were incubated at 28 °C for 72 h; the colonies growing on the plates were taken and purified by sub-culturing the isolates. Therefore, the quantitative estimation of the ACC deaminase activity was done spectrophotometrically in terms of α-ketobutyrate production at 540 nm by comparing it with the standard curve of α-ketobutyrate, which ranged from 0.1 to 1.0 μmol [46]. The protein estimation was done as per the Bradford assay [47]. One unit of the ACC deaminase activity was expressed as the amount of α-ketobutyrate liberated in nmol per milligram of cellular protein per hour.