Professor Charles Maynard insists that the Darling 4 is far from a “Frankentree,” despite its genetic modifications.
For over 22 years, Maynard and Bill Powell, co-directors of the College of Environmental Science and Forestry's American Chestnut Research and Restoration Center, have attempted to engineer American chestnut trees with resistance to a specific blight that has left the species almost completely extinct by the start of the 20th century. One group of trees, all clones of a cell referred to as "Darling 4," show strong resistance to the fungus.
Many animals, including turkeys, deer, squirrels, bears and several types of birds, relied on the American chestnut to build fat for the winter before the blight struck. Once the chestnut started dwindling in numbers, these animals resorted to eating oak tree products, a much less nutritious option.
“I think it was proven by nature that when the American chestnut went down to the blight, the oak tree wasn’t able to carry [many species] over,” said Herb Darling, president of the New York chapter of the American Chestnut Foundation. “It was not too long before the turkey population decreased,” he added. The Darling 4 is named after Herb Darling and his family.
About 5 million American chestnut trees lived in American forests before the blight struck, Powell said. Millions of chestnut stumps still survive and re-sprout in nature today, but they are typically re-infected by the blight before they have a chance to flower and produce their nuts.
The blight was first discovered in the New York Botanical Garden in 1904, but it had likely been attacking trees in the United States since the late 1870s when a prominent nursery in Long Island started selling Japanese chestnut varieties, Maynard said.
The Darling 4 is one of 17 different genetically engineered cells that include a gene from wheat known as oxalate oxidase. While the gene does not kill the fungus, it does protect the tree from damage, Maynard said.
“Oxalate oxidase doesn’t harm the [fungus], but it will starve it,” explained Maynard. “It deactivates the acid that kills the plant, forcing the fungus to go back to being a bark parasite on the surface of the bark.”
The blight enters chestnut trees through small wounds before colonizing and attacking the rest of the tree. As it penetrates deeper, the oxalic acid cuts off circulation to the stem and slowly kills the tree branch by branch.
Maynard seemed proud to share the Darling 4’s results, but he believes work still needs to be done. “We’ve got solid data that Darling 4 is going to be resistant,” he said. “We just don’t think it’s going to be resistant enough to go out in the field.”
Cankers in non-transgenic American chestnut trees are usually 11,500 mm after 15 weeks of infection. In the trees with Darling 4, cankers were only about 2,750 mm after the same stretch of time. Before he is confident the plant will survive in nature, Maynard said cankers in American chestnut trees should be comparable to the naturally-resistant Chinese chestnut tree, which only gets cankers of about 1450 mm.
A new, yet-unnamed event, or genetically engineered cell, being cultivated could yield the results Maynard and Powell are looking for. It shows fewer early disease symptoms than the Chinese chestnut.
Field trees with this particular event are scheduled to be planted across New York in 2013, but it will take another three years from that point before more concrete studies can be conducted.
The ultimate goal of the Research and Restoration Center is to plant thousands of fully blight-resistant trees in American forests across the country that will slowly repopulate the forests. Powell estimates it will take about 100 years from the time the first tree is planted outside of a research station to the time the forests are fully repopulated.
Organizations such as the U.S. Department of Agriculture, the Environmental Protection Agency and the Food and Drug Administration must determine that the trees are not radically different from naturally occurring chestnut trees, other than their resistance to the blight, before a single transgenic tree can be planted in a natural habitat. This process can take up to five years.
Powell insisted the blight resistance is the only difference from naturally-occurring trees. “There are about 45,000 genes in a chestnut tree, and we are only adding one to three,” he said.
The first blight-resistant trees will be planted in public parks, private landowner properties and historic sites such as Gettysburg, said Powell.
“It’s unrealistic to think we can plant enough trees to restore the ecosystem to how it used to be without letting them multiply on their own,” said Powell. “Instead, we can plant them in memorable places in order to recreate what the scene looked like in the past, such as during the Civil War.”
There are currently 967 total trees in production at SUNY-ESF’s greenhouses and in their tissue culture labs. It takes about two years for a tree to grow from a cluster of cells in a petri dish to something large enough to plant in the ground. There are about an additional 600 transgenic trees planted in the ground at research stations across New York state, but Powell expects this number to exceed 1,000 by the spring of 2013.
Powell and Maynard hope to reach the point where they can produce 1,000 new trees a year. To accomplish this, the chestnut team changed the genetic transformation process to one that will shorten the process by three to six months, said Linda McGuigan, the director of SUNY-ESF’s tissue culture lab.
The team now uses a liquid medium called a bioreactor to kill off any non-transformed cells. The bioreactor exposes all non-transformed cells to an antibiotic at once by flooding the system every four hours.
In the past, they used what is known as a semi-solid medium, but this only exposed one side of the plant to the antibiotic at any given time. “It was kind of like a jello with plant nutrients,” said McGuigan. “It wouldn’t be something a person would want to eat, but plants love it!"