Screening, Identification Of Endofugus From Taxus Mairei And Preliminary Study On Coupling Cultivation

In order to study the relation between endofungi and their host, more than 1,000 endofungi were separated from Taxus chinensis var. mairei and were preliminarily classified identified for study on the relation of the endofungi and their host. These endofungi were mostly distributed as follow: Glomerella sp.,Botryoderma sp.,Botrytis sp.,Cephalosporium sp.,Penicillium sp.,Aspergillus sp.,Verticillium sp.,Amblyosporium sp.,Trichoderma sp.,Gilmastix sp.,Hyalodendron sp.,Phomopsis sp.,Gliomastix sp.,Fusarium sp.,Cytosporira sp.,Cytosporella sp.,Phoma sp.,Colletotrichum sp.,Eurotium sp.,Aspergillus sp.,Stroptothrix sp.,Periconia sp.,Alternaria sp.,Vlocladium sp.,Oidiodendron sp.,Basipetosrium sp.. And Fusarium sp. was the preponderant.Among which, 9 paclitaxel-producing endofungi were screened and strain Y1117 was the highest in the taxol productivity. Systematic identification for strain Y1117 were carried out including productivity, morphological and molecular biology characters, as 18S rDNA, Internal transcribed spacer,β-tubulin gene. It was confirmed that the strain Y1117 belonged to genus Fusarium as a new species and we named it Fusarium mairei cheng L. et Tao WY. The strain has been applied for patent (application number: 200510038424.8) and the patent has been published (publish number: CN1670180A).High input screening was studied on strain isolation and improvement and the better condition for isolation gained was obtained: microorganism was incubated on cellophane plate for 5 d. The mycelium was collected and was vacuum dried in 50°C for 5 h, followed by extraction with mixed solvent consisted of ethanol and chloroform (1:1) for 2 h. The extration was repeated 3 times and the extraction product was examined with HPLC/MS.The examination methods for the paclitaxel produced in the course of endofungus cultivation were validated and improved. It was shown that HPLC/MS was better one, not only it could be used to assay paclitaxel in the mycelium or broth rapidly and credibly, but also it could eliminate the influence induced by other configuration analogs.F. mairei and T. cuspidata cell were cultivated in the coupling bioreactor with self-determination knowledge property right. It was shown that there was some increase of the paclitaxel output of plant cell and microorganism cell in the cultivation, and there was some change in growth process and metabolization characters. Followed by preliminarily analysis for the above phenomena and the base was established for further study on the determinate effect substance and symbiotic principle between the endofungi and their plant host.