Editing DTH8 Gene Using CRISPR/Cas9 Technology to Improve Heading Date of Rice 99-25

doi:10.7668/hbnxb.20191109

Abstract

Abstract: CRISPR/Cas9 gene editing technology has become an important means for rice breeding. For cuttivating early maturity, high yield and high quality germplasm resources,Xiangnuo 99-25 which is a late-maturing japonica rice with excellent agronomic traits was used as the test material. The CRISPR/Cas9 dual-target expression vector was constructed to edit the heading gene DTH8 and transformed into Agrobacterium EHA105. EHA105 was transformed into rice callus via Agrobacterium-mediated transformation method, and 30 T₀ transgenic seedlings were obtained. All the T₀ seedlings were detected by PCR using hygromycin specific primer. Among them, 29 strains were detected as positive plants and the positive rate was 96.7%. Specific primers were designed to amplify 600 bp upstream/downstream of the target site in the 29 positive seedlings. Sequence results showed that 7 seedlings had mutations, such as base substitution and insertion around the second target site. The heading date of DTH8-2,DTH8-5, DTH8-9,DTH8-10,DTH8-17,DTH8-21 in these 7 seedlings were detected early heading date compared with Xiangnuo 99-25. And DTH8 expression in DTH8-2, DTH8-9, DTH8-17 were down-regulated compared with Xiangnuo 99-25. CRISPR/Cas9 gene editing technology was successfully used to edit DTH8 gene in Xiangnuo 99-25. The DTH8 mutant rice with early heading stage was obtained, which provided germplasm resources for cultivating early maturing, high-yield and high-quality rice varieties.

Key words: Rice, Heading stage, Gene editing, DTH8, CRISPR/Cas9

CLC Number:

S511.03
Q78

ZHANG Hao, LIU Xu, XUAN Ning, ZHANG Hua, GAO Ruiyu, ZHAO Qianqian, YAO Fangyin. Editing DTH8 Gene Using CRISPR/Cas9 Technology to Improve Heading Date of Rice 99-25[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2020, 35(6): 58-66. doi: 10.7668/hbnxb.20191109.

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