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利用CP培养基培养黑暗链霉菌 (Streptomycestenebrarius) 990 4,采用二级培养即首先在37℃培养 48h ,然后按 10 %的转种量转种于新鲜的培养基中 ,同时补加 2 %甘氨酸 ,2 8℃培养 2 0h ,收获的菌丝体对溶菌酶敏感。在适宜的酶解条件下可形成 4 6 2× 10 9/mL原生质体 ,再生率为18%。利用经修饰的质粒DNA转化冻存的原生质体获得成功 ,转化率为 10 3～ 10 4 / μgDNA。
The Streptomyces tenebrarius 9904 was cultured in CP medium. In the secondary culture, the cells were first cultured at 37 ° C for 48h and then transplanted into fresh medium at the rate of 10% while supplemented with 2% glycine, 2 8 ℃ cultured 20h harvested mycelia sensitive to lysozyme. Under suitable conditions for enzymatic hydrolysis, 462 × 10 9 / mL protoplasts were formed with a regeneration rate of 18%. Successful transformation of cryopreserved protoplasts with the modified plasmid DNA resulted in a conversion of 10 3 to 10 4 / μg DNA.