Maize Resources Institute, Jilin Academy of Agricultural Sciences, Gongzhuling 136100, China.
Key Laboratory of Agricultural Biotechnology, Jilin Academy of Agricultural Sciences, Changchun 130033, China.
Int J Mol Sci. 2022 May 20;23(10):5755. doi: 10.3390/ijms23105755.
CRISPR/Cas system has developed a new technology to modify target genes. In this study, CasΦ2 is a newly Cas protein that we used for genome modification in Arabidopsis and tobacco. and of marker genes were chosen for targeting. CasΦ2 has the function to cleave pre-crRNA. In the presence of 10 mM Mg irons concentration, sgRNA3 type guided CasΦ2 to edit target gene and generate mutation, and a mutant seedling of gene with an expected male sterile phenotype was obtained. In the process of tobacco transformation, the gene editing activity of CasΦ2 can be activated by 100 nM Mg irons concentration, and sgRNA1 type guided CasΦ2 to edit target gene. Mutant seedlings of gene with an expected albino were obtained. The results indicate that CasΦ2 can effectively edit target genes under the guidance of different sgRNA type in the presence of Mg ions. Together, our results verify that the CRISPR/CasΦ2 system is an effective and precise tool for genome editing in plants.
CRISPR/Cas 系统开发了一种新的技术来修饰靶基因。在本研究中，CasΦ2 是一种新的 Cas 蛋白，我们将其用于拟南芥和烟草的基因组修饰。选择了 和 两种标记基因进行靶向编辑。CasΦ2 具有切割前 crRNA 的功能。在 10mM Mg 离子浓度存在下，sgRNA3 型指导 CasΦ2 编辑靶基因并产生突变，获得了具有预期雄性不育表型的 基因的突变体幼苗。在烟草转化过程中，100nM Mg 离子浓度可激活 CasΦ2 的基因编辑活性，sgRNA1 型指导 CasΦ2 编辑靶基因。获得了具有预期白化表型的 基因的突变体幼苗。结果表明，CasΦ2 在 Mg 离子存在下，在不同 sgRNA 类型的指导下，可以有效地编辑靶基因。总之，我们的结果验证了 CRISPR/CasΦ2 系统是一种在植物中进行基因组编辑的有效和精确的工具。
