Establishment of an efficient transformation protocol and its application in marine-derived Bacillus strain
Establishment of an efficient transformation protocol and its application in marine-derived Bacillus strain
复制标题
高效转化方案的建立及其在海洋芽孢杆菌菌株中的应用
DOI：
10.1007/s11427-014-4632-3
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发表时间：
2014-06-01
影响因子：
9.1
通讯作者：
Li WenLi
中科院分区：
文献类型：
--
作者：
Liu Yang;Zheng Hua;Li WenLi
Marine-derived Bacillus strains have been proved to be a very promising source for natural product leads. However, transformation of environmental strains is much more difficult than that of domesticated strains. Here, we report the development of an efficient and robust electroporation-based transformation system for marine-derived Bacillus marinus B-9987, which is a macrolactin antibiotics producer and a very promising biological control agent against fungal plant diseases. The transformation efficiency was greatly enhanced 10(3)-fold by using unmethylated plasmid to bypass modification-restriction barrier, and using glycine betaine to protect cells from electrical damages during electroporation. Addition of HEPES and 2 mmol L(-1) MgCl2 further improved the efficiency by additional 2-fold, with a maximum value of 7.1×10(4) cfu/μg pHT3101. To demonstrate the feasibility and efficiency of the protocol, a green fluorescent protein reporter system was constructed; furthermore, phosphopantetheinyl transferase gene sfp, which is essential to the biosynthesis of polyketides and nonribosomal peptides, was overexpressed in B-9987, leading to increased production of macrolactin A by about 1.6-fold. In addition, this protocol is also applicable to marine-derived Bacillus licheniforms EI-34-6, indicating it could be a reference for other undomesticated Bacillus strains. To our knowledge, this is the first report regarding the transformation of marine-derived Bacillus strain.
