Anthocyanin, a novel and user-friendly reporter for convenient, non-destructive, low cost, directly visual selection of transgenic hairy roots in the study of rhizobia-legume symbiosis.

BACKGROUND

-mediated hairy root transformation provides a powerful tool for investigating the functions of plant genes involved in rhizobia-legume symbiosis. However, in the traditional identification methods of transgenic hairy roots based on reporter genes, an expensive chemical substrate or equipment is required.

RESULTS

Here, we report a novel, low cost, and robust reporter for convenient, non-destructive, and directly visual selection of transgenic hairy roots by naked eye, which can be used in the study of rhizobia-legume symbiosis. The reporter gene in , encoding an R2R3 type MYB transcription factor, was ectopically expressed in hairy roots-mediated by . , which induced purple/red colored anthocyanin accumulation in crop species like soybean ( (L.) Merr.) and two model legume species, and . Transgenic hairy roots of legumes containing anthocyanin can establish effective symbiosis with rhizobia. We also demonstrated the reliability of as a reporter gene by CRISPR/Cas9-targeted mutagenesis of the soybean resistance to nodulation gene in the soybean PI377578 (Nod-) inoculated with USDA193. Without exception, mature nitrogen-fixation nodules, were formed on purple transgenic hairy roots containing anthocyanin.

CONCLUSIONS

Anthocyanin is a reliable, user-friendly, convenient, non-destructive, low cost, directly visual reporter for studying symbiotic nitrogen-fixing nodule development and could be widely applied in broad leguminous plants.