CRISPR-Cas9多靶点载体构建及高效定向编辑水稻苯达松抗性基因CYP81A

发布时间：2018-04-09 07:25

  本文选题：苯达松　切入点：CRISPR-Cas　出处：《基因组学与应用生物学》2017年08期

【摘要】：水稻CYP81A6基因(Bel)编码的细胞色素P450单加氧酶对苯达松和磺酰脲类除草剂具有抗性,该基因的隐性纯合突变体(bel)表现为苯达松敏感致死性,可作为植物化学致死标记,在水稻杂交制种中具有重要的应用价值。本研究在CYP81A6基因的首个外显子中设计两个靶位点,构建了含多靶点的CRISPR-Cas9载体,通过根癌农杆菌介导法转化粳稻中花11,获得了56株转基因T0代阳性植株,提取其中9株基因组DNA进行靶位点测序检测,发现基因敲除突变率为77.8%,其中纯合突变率高达71.4%,对纯合突变体水稻喷施苯达松,均表现出对苯达松敏感致死。为应用CRISPR-Cas9技术培育苯达松敏感致死水稻不育系和对其他水稻基因进行编辑提供理论依据。
[Abstract]:The cytochrome P450 monooxygenase encoded by CYP81A6 gene of rice is resistant to bentazone and sulfonylurea herbicides. The recessive homozygous mutant of this gene is bentazone sensitive to death and can be used as a chemical lethal marker in plants.It has important application value in hybrid seed production of rice.In this study, two targets were designed in the first exon of CYP81A6 gene, and a CRISPR-Cas9 vector containing multiple targets was constructed. By Agrobacterium tumefaciens mediated transformation of Japonica rice Zhonghua 11, 56 transgenic T0 positive plants were obtained.The gene knockout rate was 77.8%, and the homozygous mutation rate was 71.4%. Bentazon was sprayed on the homozygous mutant rice, all of which showed sensitivity to bentazone.It provides theoretical basis for the cultivation of Bentazon sensitive lethal rice sterile lines and the editing of other rice genes by using CRISPR-Cas9 technique.
【作者单位】： 南昌大学生命科学学院江西省分子生物学与基因工程重点实验室;
【基金】：南昌大学“创新创业训练计划”项目、南昌大学研究生创新专项资金(cx2016193) 江西省研究生创新专项资金(YC2016-S063)共同资助
【分类号】：Q943.2
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本文编号：1725425