摘要
为建立马铃薯疮痂病新致病种Streptomyces galilaeus典型菌株CPS-2的转化体系,对该菌的原生质体制备和再生条件进行了筛选。结果表明,菌株CPS-2原生质体制备的最适条件为：TSB培养基一级培养24h,改良YEME培养基（含10%蔗糖和1.0%甘氨酸）二级培养60h,溶菌酶用量为3.0g/L,37℃酶解90 min,原生质体形成量可达4.4×109个/mL;在R2YE培养基上再生率可达22.71%。采用含25%PEG1000的Tbuffer介导转化质粒pIJ702,24h后覆盖含有硫链丝菌素的营养软琼脂,4d后可筛选到转化子,每微克质粒DNA可获得102~103个转化子。
The conditions of formation and regeneration of protoplast were screened in order to construct the protoplast transformation system of potato scab pathogen Streptomyces galilaeus（strain） CPS-2.The result showed that the optimum conditions for protoplast formation was as follows： shaking culture in TSB for 24h,transferring to YEME medium（containing 10% glucose and 1.0% glycine） and continuning shaking culture for 60h.The number of protoplasts per mL could up to 4.4×109 when the lysozyme concentration was 3.0g/L and incubated at 37℃ for 90min.The regeneration rate was 22.71% on R2YE medium.Plasmid pIJ702 was used to validated the protoplasts of strain CPS-2.Using T buffer containing 25% PEG 1000,102-103 transformants per μg plasmid DNA were obtained after four days.
出处
《河南农业科学》
CSCD
北大核心
2010年第8期71-76,共6页
Journal of Henan Agricultural Sciences
基金
国家自然科学基金项目(30700523)
国家863项目(2006AA10A211)
现代农业产业技术体系建设专项资金资助项目(NYCYTX-15)
关键词
马铃薯疮痂病菌
原生质体
再生
转化
Potato scab pathogen
Protoplast
Regeneration
Transformation
