罗田甜柿原生质体分离、培养及植株再生

Studies on the Isolation,Culture of Protoplast and Plant Regeration in Luotiantianshi Persimmon

以‘罗田甜柿’休眠芽茎尖诱导的愈伤组织为试材 ,对原生质体分离培养技术进行了研究。结果表明 :( 1)继代 10d的愈伤组织最适于分离原生质体 ;( 2 )优化的酶解体系为CPW +0 .5 %CellulaseOnorukaR 10 +0 .0 3 %PectinasefromAspergillusNiger+0 .7mol/LMannitol+1%PVP 10 ;( 3)原生质纯化方式以上浮法为好 ,10 0×g离心 6min ;( 4)琼脂糖念珠培养 5～ 6d观测到第 1次分裂 ,多为不均等分裂 ,6 0～70d形成肉眼可见的小愈伤组织 ,愈伤组织增殖到 5mm以上 ,分化成苗 ,转至生根培养基生根。

s:The conditions of isolation and culture of protoplasts from leaf primordial excised from dormant buds of adult nonastringent presimmon'Luotiantianshi'were studied.The best combination of enzymes selected for protoplast isolation from the calli subcultured 10d was CPW+0.5% Cellulas R-10+0.03% Pedinase+ 0.7mol/L Mannitol+1% PVP-10.Protoplast purification could be obtained by means of floating collection and centrifugation.Protoplast divided for the first time after 5-6d,formed multi-cell clusters and grew into visible mini-callus after 60-70d,then mini-callus was transferred onto calli agar medium for proliferation.When the calli grew to more than 5mm in diameter,they were induced to adventitious bud and rooted.