Agrobacterium tumefaciens-Mediated Transformation of Rice by Hygromycin Phosphotransferase (hptII) Gene Containing CRISPR/Cas9 Vector.

The CRISPR/Cas9 technique for rice genome engineering is gaining momentum and requires a precise gene delivery system. For rice and other crop plants, Agrobacterium tumefaciens-mediated transformation (AMT) is considered a suitable gene transformation method. The AMT for indica-type rice is a challenge because it is less efficient in tissue culture response than japonica-type rice. Here is a protocol of the AMT method that we developed for IR64 variety which has been successfully tested in other popular indica-type rice varieties. We used embryogenic calli as explant and an empty gRNA-containing CRISPR/Cas9 vector with hptII (hygromycin phosphotransferase) gene for the transformation. This technique would speed up rice genome editing via CRISPR/Cas9 technology and facilitate to achieve varied application in the future.