摘要
采用电穿孔的方法对海洋红酵母进行外源DNA的转化.通过调整参数,对影响电转化的主要因素进行探索,初步建立了载体pTEF1/Zeo-rDNA对海洋红酵母的高效电转化方法.结果表明,当采用对数生长中期的菌体制备感受态细胞、电压为900V、质粒浓度为20mg/L和0.2cm电转化杯时,转化率达到最大值,为每微克质粒DNA52个转化子.经抽样鉴定所得到的转化子均为阳性克隆.首次建立了以海洋红酵母为宿主的高效电转化体系,为外源基因在海洋红酵母中的表达奠定了基础.
Electrotransformation, a highly efficient and easy-applying technique, is used to introduce vector pTEF1 / Zeo-rDNA into Rhodotorula benthica in this work. The optimal electroporated stage for Rhodotorula benthica was at intermediate logarithmic phage. Under the conditions of voltage of 900 V and 20 mg/ L plasmid DNA and 0.2 cm cuvettes, the transformation efficiency reach its maximum at 52 transformants/μg plasmid DNA. The results showed that all of the transformants were positive clones. The experiments provided a model for highly efficient electrotransformation of Rhodotorula benthica as host cells for the first time. The research established a basis of studying biological properties of Rhodotorula benthica and its further development of genetic engineering products.
出处
《生命科学研究》
CAS
CSCD
2008年第2期136-140,共5页
Life Science Research
基金
国家自然科学基金资助项目(30560117)
关键词
海洋红酵母
高效电转化
转化率
Rhodotorula benthica
electrotransformation
transformation efficiency
