Golden-Gate compatible Magnaporthe oryzae protoplast transformation vectors

The Golden Gate cloning system uses standardised parts to facilitate the assembly of multiple transcriptional units, to ensure that future work with these genes can be carried out with ease (Patron et al., 2015 New Phytologist, v. 208, p. 13-19).Three fungal transformation vectors have been adapted from the pCB1532 vector series (Sweigard et al., 1997. Fungal Genetics Newsletter 44: 52-53). Vector pCB1532B-RFP Addgene #101854 encodes bialaphos/basta/L-phosphinothricin resistance, pCB1532H-RFP #101855 hygromycin resistance and pCB1532S-RFP #101856 sulfonylurea/chlorimuron ethyl resistance. Vectors were domesticated through removal of BsaI cloning sites. An RFP-marker was inserted. The RFP is expressed in E. coli, allowing for red-white selection of transformants. The marker is lost during the Golden Gate reaction, as it is replaced by the inserted transcriptional units. Vectors, sequence information and plasmid maps are available from Addgene https://www.addgene.org/plasmids/articles/28191792/

金门克隆系统采用标准化组件，以便利多个转录单元的组装，从而确保未来对这些基因的研究工作能够轻松进行（Patron 等人，2015 年《新植物学家》，第 208 卷，第 13-19 页）。本系统已从 pCB1532 系列矢量中衍生出三种真菌转化载体（Sweigard 等人，1997 年《真菌遗传学通讯》第 44 期：第 52-53 页）。矢量 pCB1532B-RFP（Addgene #101854）编码双草丁膦/草甘膦/L-磷酰氨基三唑抗性，pCB1532H-RFP（#101855）编码潮霉素抗性，pCB1532S-RFP（#101856）编码磺酰脲/氯磺隆乙酯抗性。通过移除 BsaI 克隆位点对矢量进行了驯化，并插入 RFP 标记。RFP 在大肠杆菌中表达，允许对转化子进行红-白选择。在金门反应过程中，标记丢失，因为它被插入的转录单元所替代。矢量、序列信息和质粒图谱可从 Addgene 获取：https://www.addgene.org/plasmids/articles/28191792/。