| The biological characteristics of Peronophthora litchii demonstrated that its sporangio- phore belonged to multideterminate. The sporangia could germinate by means of tubes, zoospores, and secondary sporangia. It could form chlamydospores after culturing for 35d. The optimum conditions for the mycelia growth were that they growed on LBA medium (pH7.0) at 24℃and under light for 20h. The optimum conditions of spores formation were that they growed on PDA medium (pH7.0) at 24℃and under light for 16h. The lethal temperature of the sporangia was 55℃, keeping for 10 min. The best germination rate of the sporangia was from the sporangia which were produced from the fungus cultured for 11 days at 24℃and pH 7. The results showed that the germination form and rate of sporangia were different when they in various temperatures, incubation times, pH values and nutrient compositions.This study has optimized preparation and regeneration of protoplasts from P. litchii. The medium was PDA, the enzyme concentrations was 15mg·mL-1 driselase, digesting time was 4h, digesting temperature was 30℃, incubating time was 4d, regeneration medium was LBA with 0.6M mannitol as osmotic stabillzer. The quantity and quality of protoplasts producted satisified the demand of protoplast-media transformation with the yield of 107 cfu·mL-1 and a regeneration frequency of 0.1% .The study had constructed a genetic transformation system of P. litchii and GFP-expression vector. Recombinant vector pHAMT35G–gfp and vector pTH209 were transformed into the protoplasts of the fungus, and resulted in transformants. After screening with selective plates, PCR amplification of targete fragment, it was definite that nptⅡand GFP had been inserted in the genome of P. litchii. |
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