摘要
为了得到耐贮藏的转基因柿,以上西早生柿叶片为外植体,通过农杆菌介导法,建立了ACC氧化酶的RNAi遗传转化体系。研究了预培养时间、农杆菌菌液浓度、侵染时间、乙酰丁香酮(AS)浓度和共培养时间等因素对基因转化的影响。结果表明:适宜的壮观霉素浓度为30 mg/L,预培养时间为3 d,用OD600值为0.3的菌液侵染10 min、AS浓度100 mg/L、共培养4 d有利于提高转化频率。经PCR分子检测与GUS组织化学染色,初步确定ACC氧化酶基因已转入上西早生柿组培苗中。
In order to get transgenic Japanese Persimmon,using young leaves of Uenishiwase as ex- plant, ACC oxidase gene was transferred into Uenishiwase, mediated by Agrobacterium tumefa-ciens. The pre-cultured time, the concentration of A. tumefaciens, the infection time, the con- centration of AS and the co- cultured time were studied. Together studied was the genetic transfor- mation systems. The results showed that the available concentration of Spectinomycin was 10 mg/L, the best pre- cultured time was 3 d, the fittest OD600 of engineering bacterial suspension was 0.3, the optimum infection time was 10 min, the co- cultured time was 4 d and AS 100 mg/L could in- crease the frequency of transformation. It was originally proved by the PCR examination and GUS assays that the ACO gene had been transferred into Uenishiwase.
出处
《河北农业大学学报》
CAS
CSCD
北大核心
2009年第2期66-70,共5页
Journal of Hebei Agricultural University
基金
河北省国际科技合作攻关项目(04395501D-3)
