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. 2023 Mar 14;14:1134653. doi: 10.3389/fpls.2023.1134653

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Copyright © 2023 Huang, Lin, Cao, Zhang, Song, Huang, Sun, Long, Liao and Zhuo

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Detection of Homozygous OsHPP04 mutants of rice. (A) Sanger sequencing results for OsHPP04 mutants in T₁ generation. The protospacer adjacent motif (PAM) sequences are shown in blue box. DSB indicate double-strand breaks. The red arrowheads indicate the location of mutations. (B) PCR-based test for identifying ‘transgene-free’ OsHPP04 mutants of rice. Cas9p (732 bp), part of the Cas9 coding sequence. sgRNA (723 bp), region ranging from the OsU6a promoter to the downstream of gRNA2. 18S rRNA (453 bp), part of the 18s rRNA sequence as a normalization control. M, DL2000 DNA marker. Nip, wild-type rice DNA. P, plant pYLCRISPR/Cas9-Pubi-H-OsHPP04-KO binary vector. Lanes 1-11 and 12-18, individual offspring of Line H4 and H6.