Co-expression of GR79 EPSPS and GAT generates highglyphosate-resistant alfalfa with low glyphosate residues

Weed competition seriously threatens the yield of alfalfa, the most important forage legume worldwide,thus generating herbicide-resistant alfalfa varieties is becoming a necessary cost-effective strategy toassist farmers for weed control. Here, we report the co-expression of plant codon-optimized forms ofGR79 EPSPS (pGR79 EPSPS) and N-acetyltransferase (pGAT) genes, in alfalfa, via Agrobacterium-mediated transformation. We established that the pGR79 EPSPS-pGAT co-expression alfalfa lines were able totolerate up to tenfold higher commercial usage of glyphosate and produced approximately ten timeslower glyphosate residues than the conventional cultivar. Our findings generate an elite herbicideresistant germplasm for alfalfa breeding and provide a promising strategy for developing high-glyphosate-resistant and low-glyphosate-residue forages.