Intron-targeted gene insertion in rice using CRISPR/Cas9: A case study of the Pi-ta gene

Publication date: Available online 5 June 2019Source: The Crop JournalAuthor(s): Yang Xu, Fangquan Wang, Zhihui Chen, Jun Wang, Wen-Qi Li, Fangjun Fan, Yajun Tao, Ling Zhao, Weigong Zhong, Qian-Hao Zhu, Jie YangAbstractIntron-targeted gene insertion strategy using CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated Cas9) has been shown to be a potential tool for crop genetic improvement by targeted mutagenesis or gene replacement of an elite allele into widely cultivated rice varieties. The rice blast resistant protein Pi-ta, differs from its susceptible counterpart, pi-ta, by a single amino acid in exon 2. To create new materials resistant to the rice blast disease, we inserted a genomic fragment containing the exon 2 and 3′ untranslated region (3′ UTR) of Pi-ta into intron 1 of pi-ta in rice materials susceptible to rice blast using the intron-targeted insertion strategy. The gene insertion frequency was 3.8%. Several novel transgene-free progeny with stably inherited homozygous insert were identified in the T1 generation, which have been crossed to rice germplasm bearing other resistance gene (R gene) for pyramiding of R genes. This work verified the feasibility of using the genome editing technology in improvement of qualitative agronomic trait in crops.

Source: The Crop Journal - June 6, 2019 Category: Food Science Source Type: research