桃褐腐病菌(Monilia fructigena)原生质体制备及再生条件

Protoplast Preparation and Regeneration of Monilia fructigena

摘要:

以桃褐腐病菌(Monilia fructigena)为供试菌株, 研究了酶系组成、液体培养基、菌龄、酶解温度、酶解时间对原生质体制备的影响, 以及等渗液、固体再生培养基、酶解时间对原生质体再生的影响。结果表明: Fries(1/2)液体培养基培养24 h, 在10 mg/mL崩溃酶 + 5 mg/mL纤维素酶 + 20 mg/mL蜗牛酶 + 10 mg/mL溶菌酶的混合酶液中28?C酶解4 h为桃褐腐病菌原生质体制备的最佳条件。采用液体再生涂布平板法, 以含Ca2+的STC为等渗液的液体培养基和含蔗糖及Ca2+的Fries(1/2)固体培养基为桃褐腐病菌原生质体再生的最佳条件。经过观察与测定, 再生菌株保持了原有的培养性状和致病性, 接种桃果实后发病率为100%。

Abstract:

The condition of protoplast preparation and regeneration of Monilia fructigena was studied in this research. The results showed that the composition of cell wall degradative enzymes, liquid medium, mycelial age, digesting temperature and time duration affected the preparation of protoplast. More proto-plast of M. fructigena were yielded when mycelia incubated in Fries(1/2) for 24 h were digested by en-zyme mixture of 10 mg/mL driselase, 20 mg/mL snailase, 5 mg/mL celluase and 10 mg/mL lysozyme for 4 h at 28?C. The protoplast regeneration rate was affected by osmotic solution, solid regeneration medium, and digesting time.The best regeneration of protoplast of M. fructigena was achieved by using STC as isotonic solution and Fries(1/2) solid medium as regeneration medium. The regenerated strain of M. fruc-tigena was same as original strain in colony morphology and pathogenicity.