Developing a flexible, high‐efficiency <i>Agrobacterium</i>‐mediated sorghum transformation system with broad application

TLDR

Sorghum, the fifth most widely planted cereal crop, is essential in arid regions but genetic improvement via transgenics has been limited by an inefficient transformation system. The study introduces a ternary vector system incorporating a pVIR accessory plasmid to enable efficient Agrobacterium‑mediated transformation of sorghum. Using this ternary vector, the authors transformed sorghum, applied the protocol to several recalcitrant African varieties, and established the first stable CRISPR/Cas9 knockouts in Tx430. The system yielded 6–29% regeneration in Tx430, produced 45–66% single‑copy backbone‑free events, and facilitated transformation of recalcitrant varieties and CRISPR/Cas9 knockouts.

Abstract

Summary Sorghum is the fifth most widely planted cereal crop in the world and is commonly cultivated in arid and semi‐arid regions such as Africa. Despite its importance as a food source, sorghum genetic improvement through transgenic approaches has been limited because of an inefficient transformation system. Here, we report a ternary vector (also known as cohabitating vector) system using a recently described pVIR accessory plasmid that facilitates efficient Agrobacterium ‐mediated transformation of sorghum. We report regeneration frequencies ranging from 6% to 29% in Tx430 using different selectable markers and single copy, backbone free ‘quality events’ ranging from 45% to 66% of the total events produced. Furthermore, we successfully applied this ternary system to develop transformation protocols for popular but recalcitrant African varieties including Macia , Malisor 84‐7 and Tegemeo . In addition, we report the use of this technology to develop the first stable CRISPR /Cas9‐mediated gene knockouts in Tx430.