Efficient CRISPR/Cas9‐mediated genome editing in sheepgrass (<i>Leymus chinensis</i>)
Zhelong Lin, Lei Chen, Shanjie Tang, Mengjie Zhao, Tong Li, Jia You, Changqing You, Boshu Li, Qinghua Zhao, Dongmei Zhang, Jianli Wang, Zhongbao Shen, Xianwei Song, Shuaibin Zhang, Xiaofeng Cao
Abstract
The lack of genome editing platforms has hampered efforts to study and improve forage crops that can be grown on lands not suited to other crops. Here, we established efficient Agrobacterium-mediated clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated nuclease 9 (Cas9) genome editing in a perennial, stress-tolerant forage grass, sheepgrass (Leymus chinensis). By screening for active single-guide RNAs (sgRNAs), accessions that regenerate well, suitable Agrobacterium strains, and optimal culture media, and co-expressing the morphogenic factor TaWOX5, we achieved 11% transformation and 5.83% editing efficiency in sheepgrass. Knocking out Teosinte Branched1 (TB1) significantly increased tiller number and biomass. This study opens avenues for studying gene function and breeding in sheepgrass.