Chloroplast and mitochondrial DNA editing in plants [post]

Jin-Soo Kim, Beum-Chang Kang, Su-Ji Bae, Seonghyun Lee, Jeong Sun Lee, Annie Kim, Hyunji Lee, Gayoung Baek, Huiyun Seo, Jihun Kim
2021 unpublished
Plant organelles, including mitochondria and chloroplasts, contain their own genomes, which encode hundreds of genes essential for respiration and photosynthesis, respectively. Gene editing in plant organelles, an unmet need for plant genetics and biotechnology, has been hampered by the lack of appropriate tools for targeting DNA in these organelles. In this study, we developed a Golden Gate cloning system, composed of 16 expression plasmids (8 for delivery of the resulting protein to
more » ... rotein to mitochondria and the other 8 for delivery to chloroplasts) and 424 TALE sub-array plasmids, to assemble DddA-derived cytosine base editor (DdCBE) plasmids and used the resulting DdCBEs to promote point mutagenesis in mitochondria and chloroplasts efficiently. Our DdCBEs induced base editing in lettuce or rapeseed calli at frequencies of up to 25% (mitochondria) and 38% (chloroplasts). We also showed DNA-free base editing in chloroplasts by delivering DdCBE mRNA to lettuce protoplasts. Furthermore, we generated lettuce calli resistant to streptomycin, an antibiotic that binds to 16S ribosomal RNA (rRNA) irreversibly, leading to inhibition of protein synthesis, by introducing a point mutation in the chloroplast 16S rRNA gene.
doi:10.21203/rs.3.rs-145710/v1 fatcat:zui566iporc4hjdsv67ias2a2m