Improved Transformation and Regeneration of <i>Indica</i> Rice: Disruption of <i>SUB1A</i> as a Test Case via CRISPR-Cas9

oleh: Yuya Liang, Sudip Biswas, Backki Kim, Julia Bailey-Serres, Endang M. Septiningsih

Format: Article
Diterbitkan: MDPI AG 2021-06-01

Deskripsi

Gene editing by use of clustered regularly interspaced short palindromic repeats (CRISPR) has become a powerful tool for crop improvement. However, a common bottleneck in the application of this approach to grain crops, including rice (<i>Oryza sativa</i>), is efficient vector delivery and calli regeneration, which can be hampered by genotype-dependent requirements for plant regeneration. Here, methods for <i>Agrobacterium</i>-mediated and biolistic transformation and regeneration of <i>indica</i> rice were optimized using CRISPR-Cas9 gene-editing of the submergence tolerance regulator <i>SUBMERGENCE 1A-1</i> gene of the cultivar Ciherang-Sub1. Callus induction and plantlet regeneration methods were optimized for embryogenic calli derived from immature embryos and mature seed-derived calli. Optimized regeneration (95%) and maximal editing efficiency (100%) were obtained from the immature embryo-derived calli. Phenotyping of T<sub>1</sub> seeds derived from the edited T<sub>0</sub> plants under submergence stress demonstrated inferior phenotype compared to their controls, which phenotypically validates the disruption of <i>SUB1A-1</i> function. The methods pave the way for rapid CRISPR-Cas9 gene editing of recalcitrant <i>indica</i> rice cultivars.