An optimized protocol for in vitro regeneration and genetic transformation of broomcorn millet

Abiotic stress has been a great challenge to global food security. To reduce its effects, breeding crops for tolerance to abiotic stresses is a promising strategy. Broomcorn millet is cultivated in arid and semiarid areas with a high degree of abiotic stress tolerance. However, due to the lack of efficient genetic transformation methods for broomcorn millet, the characterization of genes related to abiotic stress tolerance lags behind that of other crop species. Therefore, establishing efficient in vitro regeneration and genetic transformation methods for broomcorn millet is essential.

In this study, we used mature seeds of the sequenced variety 'Longmi 4' as explants and optimized its in vitro regeneration and genetic transformation methods. The optimal hormone concentrations for embryogenic callus induction medium were 2.5 mg/L 2,4-dichlorophenoxyacetic and 0.5 mg/L 6-benzylaminopurine. The optimal hormone concentrations for shoot regeneration medium were 2 mg/L 6-benzylaminopurine and 0.5 mg/L a-naphthaleneacetic acid. Additionally, the co-cultivation time was 3 days, and the optimal hygromycin concentration for putative transgenic callus selection was 20 mg/L. The transformation efficiency was 21.25% after our modification approach.

Here, we present a simple and highly efficient Agrobacterium-mediated genetic transformation protocol for broomcorn millet. Our work provides a tool for the characterization of genes related to important traits, as well as a new strategy for broomcorn millet breeding.