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More-Precise Genetic Engineering for Plants

New technology makes it possible to alter plant genes precisely and efficiently.

Genetically engineering plants is a time-intensive process. Methods currently used to deliver genetic changes are imprecise, so it’s often necessary to generate thousands of plants to find one that happens to have the desired alteration. Two papers in this week’s Nature detail the use of a genetic technology that allows scientists to target plant genomes more precisely. The method, which has previously been used in animals and in human cells, can be used to introduce a new gene, make small changes in existing genes, or block a gene from being expressed; it also makes it possible to introduce several different genetic changes into the same plant.

Gene fixing: Using a new technique, researchers altered a gene in tobacco plants to make them herbicide resistant. The plants shown on the left are able to thrive when exposed to herbicide, compared with control plants at right.

“We now have some control over the plant’s genetic code,” says Daniel Voytas, lead author of one of the papers and a geneticist at the University of Minnesota. The technique not only allows for more precise changes, but it greatly increases the efficiency of generating genetically engineered plants for use as food or fuel, or for absorbing carbon and cleaning the environment. “If you can deliver a gene to the same location every time with precision, that might change the regulatory landscape and decrease the cost of creating these transgenic plants,” he says.

Vipula Shukla, a scientist at Dow AgroSciences, who led the other study, says that for plant scientists, “all of the conventional tools that are available to us are based on methods that make random modifications to plant genomes.” These methods include using a bacterial vector to transfer DNA into a plant cell, or physically blasting DNA-coated particles into cells. DNA introduced these ways, Shukla says, can land anywhere within the plant’s genome and have unintended side effects like altering an existing gene or producing multiple copies of the gene of interest. Scientists typically generate many plants and then screen them to find the ones in which the desired change was successful.

Both of the new studies–one was led by Dow and another by an academic consortium–employed a gene-targeting technology called zinc finger nucleases–synthetic proteins that can precisely target locations in the genome and make specific genetic changes.

Zinc finger nucleases work by breaking both strands of DNA at a specific site in the genome. This double break prompts the cell’s own repair machinery to patch the rift. The machinery will often search for a piece of DNA that is similar to the damaged region to copy and paste back into the genome. By supplying a piece of DNA that contains sequences from the original gene with the desired changes–either the addition of a new gene or a change in sequence–scientists can induce the cell to change the genetic code as it repairs the break. The technology can also be used to block a gene by taking advantage of another repair mechanism in which the cell simply joins the two broken ends back together, which often deletes or inserts new DNA sequences into the repair site, resulting in DNA code that can’t be read properly.

The Dow group used the method to introduce two changes into maize, a plant that is often used for animal feed. The researchers targeted a gene involved in the production of phytates, chemicals in maize that most animals can’t digest, and used the gene as a landing pad to insert another gene that gives the plant tolerance to herbicides. At the same time, they disrupted the target gene so that the plant produces fewer phytates, which Shukla says can also accumulate as waste in water runoff from farms. The ability to “stack” desired traits in this way is not easily performed with existing technologies.

The academic group used a similar method, developed by the Zinc Finger Consortium, an international team of researchers committed to developing a publicly available platform for engineering zinc finger nucleases. Rather than add a new gene into a plant, the researchers used zinc finger nucleases to introduce an altered genetic sequence into an existing gene in tobacco plants; the protein encoded by the gene is a target of herbicides, and the alterations make the plants herbicide resistant. Voytas says that being able to make such subtle changes within a gene will give researchers a new way to study plant biology.

The method still requires generating multiple plants and screening them to find the ones that were successfully altered, but the numbers are in the tens or hundreds, rather than the thousands or tens of thousands. Shukla estimates that the technology cuts the time required to engineer a plant by about half. The method also requires the creation of zinc finger nucleases that are specific to a particular application. Shukla says that Dow is already employing its platform for creating the molecules across its internal products as well as in academic research projects, and it’s planning to license the technology for academic, commercial, and humanitarian use. Voytas says that the Zinc Finger Consortium is making its method available publicly and will be offering training sessions in the technique.

Matthew Porteus, a biochemist at the University of Texas at San Antonio, who wrote an accompanying editorial in Nature, says that the two papers are the first examples of investigators who have picked a gene of interest, designing zinc finger nucleases for that gene, and using the nucleases to create specific modifications in plants. Porteus, who has been investigating zinc finger nucleases as a method for gene therapy in humans, says that interest in zinc finger nucleases has been growing in the past few years. They are being used as a way to create precise mutations in zebra fish, and a human clinical trial is just beginning that will test the use of zinc finger nucleases to create genetic alterations in the T cells of patients with HIV, with the hope of making their cells better able to fight infection.

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