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海洋芽孢杆菌(Bacillus marinus)A72代谢产物macrolactin C对甘蔗黑穗病菌具有很强的抑制作用，但产量较低不利于推广应用。代谢工程手段是提高其产量的有效策略，而高效的遗传操作体系和关键基因是决定因素。通过对菌株培养条件、外源DNA修饰以及电脉冲强度进行优化，建立A72高效的遗传转化体系，遗传转化效率为(2.86±0.54)×104 CFU/μg DNA,比优化前提高658.3倍。结合基因组测序技术和Antismash在线平台，阐明A72聚酮化合物合成酶(Polyketide synthase, PKS)基因簇结构组成。过表达pksE基因，工程菌macrolactin C合成速率加快，比对照组提前16 h完成发酵过程，初步确认pksE基因为调控macrolactin C合成的潜在关键基因。本研究建立了A72高效的遗传转化体系，成功解析PKS基因簇并初步确认一个潜在关键基因，为后期代谢工程改造提供技术保障和潜在靶点。
Abstract：Macrolactin C, a metabolite of Bacillus marinus A72, show excellent inhibitory effect on causing pathogenic fungus Sugarcane Smut, however, the low macrolactin C yield of A72 hinders its application. Metabolic engineering is an effective strategy to improve metabolites yield, and efficient genetic operation system and core genes are determining factors. To establish the genetic transformation system of A72, factors affected the electric shock transformation efficiency including culture conditions, exogenous unmethylated DNA, and electric pulse intensity was studied. By using the optimized protocol, a transformation efficiency of(2.86±0.054)×10~4CFU/μg DNA was obtained, which was greatly enhanced 658.3-fold comparing with the control. Combined genome sequence with Antismash database, polyketide synthase(PKS) gene cluster was identified. By over-expressing pksE gene, macrolactin C synthesis of engineering strain was accelerated, almost 16 hours earlier than the control completing the fermentation process. In this study, we established the genetic transformation system of A72 successfully, the identified PKS gene cluster and potential core gene would be provided as targets for further improving macrolactins production of A72 by metabolic engineering.
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基本信息:
DOI：10.13417/j.gab.041.001235
中图分类号:Q78
引用信息:
[1]罗双宇,黄明娟,梁莉芬,等.Bacillus marinus A72遗传转化体系建立及其PKS基因簇解析[J].基因组学与应用生物学,2022,41(06):1235-1243.DOI:10.13417/j.gab.041.001235.
基金信息:
国家自然科学基金项目(31960019; 82060640); 广西自然科学基金项目(2019GXNSFBA185023)共同资助
2022-03-08
2022-03-08
2022-03-08