Targeted gene disruption of ATP synthases 6-1 and 6-2 in the mitochondrial genome of Arabidopsis thaliana by mitoTALENs.
Shin-ichi Arimura*, Hiroki Ayabe, Hajime Sugaya, Miki Okuno, Yoshiko Tamura, Yu Tsuruta, Yuta Watari, Shungo Yanase, Takaki Yamauchi, Takehiko Itoh, Atsushi Toyoda, Hideki Takanashi, Nobuhiro Tsutsumi
We recently achieved targeted disruptions of cytoplasmic male sterility (CMS)‐associated genes in the mitochondrial genomes of rice and rapeseed by using mitochondria‐targeted TALENs (mitoTALENs). It was the first report of stable and heritable targeted gene modification of plant mitochondrial genomes. Here, we attempted to use mitoTALENs to disrupt two mitochondrial genes in the model plant Arabidopsis (Arabidopsis thaliana) using three different promoters and two types of TALENs. The targets were the two isoforms of the ATP synthase subunit 6 gene, atp6‐1 and atp6‐2. Each of these genes was successfully deleted and the mitochondrial genomes were recovered in a homoplasmic state. The nuclear genome also has a copy of atp6‐1 and we were able to confirm that it was the mitochondrial gene and not the nuclear pseudo‐gene that was knocked out. Among the three mitoTALENs promoters tried, the RPS5A promoter was the most effective. Conventional mitoTALENs were more effective than single‐molecule mito‐compact TALENs. Targeted mitochondrial gene deletion was achieved by crossing as well as by floral‐dip transformation to introduce the mitoTALENs constructs into the nucleus. The gene disruptions were caused by large (kb‐size) deletions. The ends of the remaining sequences were connected to distant loci, mostly by illegitimate homologous recombinations between repeats.
- : The Plant Journal
- : doi:10.1111/tpj.15041
- : https://onlinelibrary.wiley.com/doi/10.1111/tpj.15041