We are developing new methods for introducing CRISPR molecules into rice for faster and easier genome editing.
The CRISPR-Cas9 genome editing technology holds promise for engineering improved agronomic traits. CRISPR-Cas9 delivery as ribonucleoproteins (RNP) allows genome editing with high efficiency and accuracy. However, currently available CRISPR-Cas9 RNP delivery approaches for crop species require tissue culture, which is a lengthy, labor-intensive process prone to random somatic mutations. Furthermore, many crop species are recalcitrant to regeneration through tissue culture, further limiting applications of CRISPR-Cas9 technology in agriculture. Here we propose to develop a novel tissue culture-free CRISPR-Cas9 RNP delivery system in the model crop species, rice. We will optimize biolistic delivery of pre-assembled Cas9-sgRNA RNPs into rice shoot apical meristem (SAM) and establish tissue culture-free plant regeneration. We will deliver the RNPs with and without repair templates to generate multiple types of genome edits. We will evaluate the efficiency and accuracy of editing through our approach and compare with those of tissue culture-dependent approaches.
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