Isolation of Endophytes from Plant Samples

Isolation of endophytes from the 43 plant samples was carried out as described by Strobel et al., [4 (link)] but with minor modifications. Plant samples, which included leaves, stems, roots, rhizomes, flowers, fruits and bark, were washed under running tap water for 10 min followed by immersion in 70% EtOH for 1 min and in NaOCl (2.5% - 5.25%) for 3 min, drained and immersed in 70% EtOH again for 30 sec. Finally, the samples were rinsed with sterile d.H₂O. Each plant sample was cut aseptically into 1 cm long segments. The cut surfaces of the segments were placed on petri dishes containing potato dextrose agar (PDA) (Oxoid) supplemented with chlortetracycline HCL (50 μg/ml, Sigma) and streptomycin sulphate (250 μg/ml, Sigma) at 28°C. Pure cultures were then transferred to PDA plates free of antibiotics and maintained in the culture collection of the Collaborative Drug Discovery Research (CDDR) Group, UiTM, Malaysia. For investigations of biological activity, the endophytes were cultivated for 14 days on PDA plates at 28°C.