枯草芽孢杆菌NCD-2菌株的高效电击转化

摘要

Transformation efficiency of Bacillus subtilis stain NCD-2 was compared when the transformation was performed by using 5 different plasmids with various sizes and replication origins.Results showed that the replication origin of plasmid had a significant effect on the transformation efficiency,but not the plasmid size.For example,plasmid pNW33N carried the replication from B.coagulans showed the highest transformation efficiency by 3.53×104 cfu·μg-1 DNA.Other parameters affecting the transformation efficiency,including electric resistances,electric field strengths and cell growth phases,were also optimized with plasmid pNW33N.The result revealed that the highest transformation efficiency of 6.07×104 cfu·μg-1 DNA in strain NCD-2 could be obtained under the optimized condition,the cells of strain NCD-2 electroporated at 14.0 kV·cm-1 and 200 Ω after incubated at 150 r·min-1 and 37 ℃ in growth medium for 3.5 h.%本研究对影响枯草芽孢杆菌Bacillus subtilisNCD-2菌株遗传转化的因素进行摸索并对其转化条件进行优化,建立了该菌株的高效电击转化体系。通过比较5个不同大小和携带不同来源复制子的质粒对其转化效率的影响,发现质粒大小与NCD-2菌株的电击转化效率之间没有相关性;而携带来源于Bacillus coagu-lans复制子的质粒pNW33N时,NCD-2菌株的电击转化效率最高,达到3.53×104cfu.μg-1DNA,说明复制子来源是影响转化效率的一个重要因素。采用pNW33N质粒在不同电阻、电场