利用CRISPR/Cas9创造早熟香味水稻

Breeding of Early-maturatity and Fragrant Rice via CRISPR/Cas9 Mediated Genome Editing

黑龙江省是我国最大的水稻种植区,近年来水稻种植面积不断增加,为加快高纬度、低积温的地区品种选育进程,本研究利用CRISPR/Cas9基因编辑技术对第二积温带主栽品种绥粳14的抽穗期基因Hd2、香味基因Badh2进行编辑。通过T_1代的分子鉴定,Hd2和Badh2基因被敲除而且基因型纯合。田间鉴定表明,转基因抗性标记也被成功剔除的两个株系,绥粳14-d1和绥粳14-d2,比绥粳14提早13天左右。利用咀嚼法、氢氧化钾浸泡法和气相色谱-质谱联用技术(GCMS)测定香味,结果显示绥粳14-d1和绥粳14-d2中香味物质2-乙酰-1-吡咯啉(2-AP)含量显著高于绥粳14。综合以上结果,本研究利用基因编辑技术,成功获得了改良的抽穗期缩短香米品种,为早熟、优质水稻品种的选育提供了理论指导,加速了育种进程。

Heilongjiang province is the largest rice cultivation region, and the growth area increased dramatically over the past years. To develop the variety with both early - maturatity and high quality, which can be grown in higher latitude region with lower tem- perature, in this study, through CRISPR/Cas9 mediated genome editing, we knocked out the flowering time gene Hal2 and fragrance gene Badh2 in Suijinglg via, a major elite cultivar in the second accumulated temperature zone. Two edited lines were generated and defined as Suijing14 - dl and Suijing14 - d2. At T, generation, through molecular characterization, representative Suijing14 - dl and Suijing14 - d2 seedlings were chosen in which both Hd2 and Badh2 were mutated and are homozygous, and the Hygromycin resistance gene was omitted.. Field statistical analysis demonstrated that the heading date of Suijing14 - dl and Suijing14 - d2 was shortened a- round 13days compared with Suijing14. Seeds aromas were preliminarily analyzed by chewing and potassium hydroxide extraction as- say. Gas Chromatography - Mass Spectrometry （ GC - MS） assay was further performed to quantify the aroma compound, 2 - acetyl - 1 - pyrrolin （2 - AP） in mature seeds, and it was found that the 2 - AP content in Suijing14 - dl and Suijing14 - d2 was significantly higher than that in Suijing14. In summary, we successfully generated early- maturatity and fragrant rice variety via CRISPR/Cas9 me- diated genome editing, which will markedly facilitate the local breeding program.