利用CRISPR/Cas9技术创制早熟甘蓝型油菜材料

Creation of Early Flowering Rapeseed Using CRISPR/Cas9 Technology

【目的】对甘蓝型油菜浙油50进行基因编辑,创制油菜早熟材料,为培育早熟油菜新品种奠定基础。【方法】BnFRI和BnFLC基因是控制油菜开花的两个关键基因。利用CRISPR/Cas9基因编辑技术对甘蓝型油菜浙油50的开花关键基因BnFRI和BnFLC基因进行编辑,农杆菌侵染甘蓝型油菜下胚轴获得具有明显早熟性状的油菜株系。【结果】对T0代转基因株系靶位点序列检测和突变类型分析,结果表明BnFLC和BnFRI基因分别有一个位点被成功编辑,编辑类型为单碱基的插入和7个碱基片段的缺失。该植株明显比正常株系提早抽薹和开花,对突变早熟单株的T1代植株进行测序检测分析,发现突变类型可稳定遗传。【结论】BnFLC和BnFRI同源基因敲除导致功能突变,可能是甘蓝型油菜提早开花的重要原因。

【Objective】The gene of Zhe You 50 of Brassica napus was edited to create early maturing materials of Brassica napus,and breed early maturing varieties of Brassica napus.【Method】BnFRI and BnFLC genes are two key genes to control the flowering of Brassica napus. In this study,The CISPR/Cas9 technique was used to edit the BnFRI and BnFLC genes in Zhe You 50 of Brassica napus. After Agrobacterium tumefaciens infecting the hypocotyl of Brassica napus,we obtained the lines with obvious precocious traits.【Result】Sequencing of T0 transgenic lines showed that BnFLC and BnFRI gene was successfully edited,and the editing type was single base insertion and seven base fragments deletion. The transgenic lines obviously bolting and flowering earlier than normal lines. T1 generation transgenic line sequencing shows that mutation can be stably inherited to the next generation.【Conclusion】The functional mutation caused by the knockout of BnFLC and BnFRI homologous genes may be an important reason for the early flowering of Brassica napus.