Taxol-producing endophytes were isolated from different plant tissues (bark, stem, and needle) of Taxus sp. collected from Shimla, Himachal Pradesh (India). Erlenmeyer flask containing 100 mL modified S7 liquid medium was seeded with 4.79 × 10⁴ spores per mL of the fungal isolate TPF-06 and incubated at 25 ℃ for 21 days with agitation speed at 150 rpm in an incubator shaker.

Bark samples (1 × 3 cm) were harvested from the stem of relatively young T. baccata subsp. wallichiana from Bhadrewah(district Doda, Jammu & Kashmir, India) Pieces of agar block containing the mycelia mats from a 7-10-day-old culture plate of TBPJ-B were transferred to 100 ml of sterilized S-7 media and incubated at 25-28 &#8451 on a rotary shaker for 5 days. This culture was used as seed culture for taxol production, where 10-20 ml of seed culture was transferred to a 2 L Erlenmeyer flask containing 500 ml of sterilized S-7 mediumand incubated at 25-28 &#8451 for 21 days in the dark as the stationary culture.

This fungus was isolated from the surface-disinfected (80% ethanol) inner bark of one tree in an old-growth cedar forest in northern Montana. Taxomyces andreanae was cultured bytransferring hyphal tips from water agar, on which bark pieces had been cultured, onto a modified-mycological agar( S-7 medium consists of 1 g of glucose, 3 g of fructose, 6 g of sucrose, 1 g of Na⁺-acetate, 1 g of soytone, 1 mg of thiamine, 1 mg of biotin, 1 mg of pyridoxal, 1 mg of Ca²⁺-pantothenate, 3.6 of mg MgSO₄, 6.5 mg of CaNO₃, 1 mg of Cu(N03)2, 2.5 mg of ZnSO₄, 5 mg of MnCI₂, 2 mg of FeCI₃, 5 mg of phenylalanine, 100 mg of Na⁺-benzoate, and 1 ml of 1 M KH₂PO₄ buffer (pH 6.8) per liter. Sugar ratio is identical to that occurring in the inner bark of Pacific yew. Modified mycological agar consists of 10 g of bacto-soytone, 40 g of glucose, 15 g of bacto-agar, 1 g of Na⁺-acetate, and 50 mg of sodium benzoate per liter. ). The mycelium was then successively transferred to eliminate traces of taxol or other taxanes carried over from the original tree source.Transfers of small agar plugs to broth cultures were made from mycelia that hadgrown 3 to 7 days. After this period, mycelia appeared to go into a quiescent state.Taxomyces andreanae was stored in water at 4 ℃ and grown on a semi-defined culture medium. The conidia of T. andreanae do not germinate, therefore we transferred pieces (0.5 by 0.5 cm) of agar block containing the mycelial mats to sterilized S-7 medium. Optimum conditions were as a still culture, at 25 ℃ , with a surface-to-volume ratio of 1.3 (cm²:ml). After 21 days of incubation, the culture was filtered through cheesecloth. Chlorocholine chloride (at 1 mg/ml) in the medium abolished taxol production.