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发明公开
CN110079530A 一种源自布氏乳杆菌的基因编辑工具及其制备方法和应用 无效 - 驳回

一种源自布氏乳杆菌的基因编辑工具及其制备方法和应用
摘要：
本发明提供了一种源自布氏乳杆菌的基因编辑工具及其制备方法和应用，属于基因编辑技术领域，所述源自布氏乳杆菌的基因编辑工具包括pcDNA3.1-LbCas9质粒和pLbCas9-sgRNA质粒。所述应用包括以下步骤：确定靶标序列，设计单链寡核苷酸对；退火获得双链DNA片段；连接到pLbCas9-sgRNA质粒中获得靶标序列sgRNA表达载体；靶标序列sgRNA表达载体与pcDNA3.1-LbCas9质粒共转染细胞后培养。所述基因编辑工具所识别的3’-端PAM序列是5’-NNAAAA-3’(不同于传统的5’-NGG-3’)，扩展了可以编辑的基因位点的范围。
摘要（英）：

The invention provides a gene editing tool originated from lactobacillus buchneri and a preparation method and application thereof, and belongs to the technical field of gene editing. The gene editingtool originated from the lactobacillus buchneri comprises pcDNA3.1-LbCas9 plasmid and pLbCas9-sgRNA plasmid. The application comprises the following steps: determining a target sequence, and designing single-stranded oligonucleotide pairs; annealing to obtain a double-stranded DNA fragment; connecting the double-stranded DNA fragment into the pLbCas9-sgRNA plasmid to obtain a target sequence sgRNA expression vector; co-transfecting cells through the target sequence sgRNA expression vector and the pcDNA3.1-LbCas9 plasmid and culturing. 3'-end PAM sequence recognized by the gene editing tool is5'-NNAAAA-3' (different from traditional 5'-NGG-3'), and the range of editable gene locus is expanded.

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