Herbicide tolerance heats up

Gene shuffling technique could deal a blow to agbio establishment.

Two US seed companies may soon challenge the dominance of US agrochemical company Monsanto in the genetically modified (GM) crop market. Monsanto is currently the leading global producer of GM crops, mainly as a result of its Roundup Ready range of glyphosate-tolerant crops. In the US, around 80 per cent of planted soybeans and cotton are now Roundup Ready varieties. Now, Maxygen and Pioneer Hi-Bred are muscling in on this market and have recently developed a novel method for conferring glyphosate-tolerance to plants.

Glyphosate is a popular broad-spectrum herbicide which inhibits the plant enzyme enolpyruvyl-shikimate-3-phospate synthase (EPSPS), which is involved in the synthesis of aromatic amino acids. Monsanto’s glyphosate-tolerant GM crops contain a gene that codes for an EPSPS-related enzyme that is not affected by glyphosate.

Other herbicide-tolerant crops, such as those tolerant to glufosinate, have been genetically modified to produce enzymes that detoxify the herbicide. The Maxygen and Pioneer Hi-Bred researchers thus wondered whether they could develop a glyphosate-tolerance trait based on the same mechanism.

The researchers looked for an enzyme capable of carrying out N-acetylation of glyphosate to produce the non-herbicidal compound N-acetylglyphosate. Using mass spectrometry, they screened several bacterial species to see if they would produce N-acetylglyphosate when incubated with glyphosate.

The best performing species was a strain of Bacillus licheniformis and the researchers identified three genes that coded for glyphosate N-acetyltransferase (GAT) enzymes. Sadly, although these enzymes could acetylate glyphosate, they were not particularly efficient.

To improve efficiency, the researchers used a technology developed by Maxygen called MolecularBreeding to randomly shuffle fragments of the three gat genes to create new variants. The team tested the enzymes coded by these variants for their ability to acetylate glyphosate, and chose the best performers as the basis for a new round of shuffling. After 11 rounds, they had created variants with a 10 000-fold improvement in the ability to acetylate glyphosate.

Introducing these 11th round variants into maize produced plants that could tolerate up to six times the usual concentration of glyphosate. Field trials of this GM maize are now underway, although it will be at least five years before it reaches the market.

Jon Evans