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Mapping the maize genome is subject of new $13 million program

Since the turn of the century, the average corn yield per acre in the United States has grown fivefold.

"If you applied this same improvement to cars," says Virginia Walbot, "then the Model-T that cost $1,000 and got 30 miles per gallon would have been replaced by a car that still costs $1,000 but gets 130 miles per gallon."

Walbot, professor of biological sciences, is the principal investigator on a $12.5 million genome research project just announced by the National Science Foundation (NSF) that is designed to keep corn yields on a continuing upward curve into the next millennium. The project has the ambitious goal of defining and sequencing all the genes of maize, about the same size as the human genome.

"To corn geneticists this is like a dream come true," Walbot says. "Five years ago we didn't think that this was even possible."

The project is part of a larger five-year, $85 million Plant Genome Research Program that will "contribute to a better understanding at the genome level of the inner workings of all plants, including economically important crops like maize, soybean, tomato and cotton," according to the NSF.

The new corn project not only holds the promise of continued increases in crop yields, but the researchers have proposed an approach that, if it proves successful, could have a significant impact on the entire field of plant genetics. Instead of first mapping the entire genome of a given species and then determining the function of individual genes, as has been done in the past, the corn geneticists have suggested a technique that will simultaneously identify genes and provide a first step in defining their function.

"There is no reason why our technique could not be applied to any plant species," Walbot says.

The purpose of genome mapping is to provide a detailed map of the heredity of different species. The DNA within the genome provides a biochemical blueprint that determines in large part how an individual plant or animal will grow.

The terms "mapping" or "sequencing" mean determining the chemical identity and position of each tiny link in the genetic chain. Genes consist of chemical units, called base pairs, that link together to form the chain of DNA, which is wound up into chromosomes. Individual genes are responsible for specific characteristics, such as height or eye color.

Learning where all the genes are, what they do and how they work together is considered an important key to understanding how different species grow. In the case of plants, scientists hope that identifying the genes for economically important traits will allow them to produce improved crop varieties.

Scientists use two basic approaches for such such genome mapping. One is the brute-force approach of sequencing the entire genome. In the case of complex organisms, like humans and corn, the genome contains billions of basic units. So sequencing it in its entirety is both costly and time consuming. It also may be unnecessary. Scientists think that there is a lot that is, of "junk" DNA in the genome, segments that are not "expressed," they do not play a role in the growth process.

That leads to the second approach, called Expressed Sequence Tags, or EST. These tags identify genes that are actually being expressed. Although it avoids the junk DNA, the EST approach has a basic limitation as well. Walbot calls it the giant sock drawer sampling problem. Suppose you had a drawer that contained millions of socks: most of them white, some of them blue, and only a very few of them red. To find a red one, forced to dig through a large number of white and blue socks. Similarly, the EST approach tends to find genes that are expressed in a large number of places. So it is useful for finding about half the genes in a large genome, she says.

The maize geneticists will try a different approach based on mobile genetic elements, called transposons. Transposons can insert themselves at many different places on a genome. Research on these jumping gene sequences has played a major role in maize research. In the process, scientists have discovered a transposon, called "Mutator," that inserts predominantly into active genes. Stanford graduate student Manish Raizada altered this transposon by inserting a sequence of bacterial DNA that allows scientists to remove the tagged gene and "immortalize" it in a bacterial culture of E. Coli. They call the genetically engineered transposon "RescueMu." This approach avoids EST's sampling problem because it is equally as likely to insert into a gene that is rarely expressed as it is one that is frequently expressed.

"We can turn huge cornfields into a few plates of bacteria," Walbot says.

The experimental plan consists of planting 21 plots consisting of 2,304 plants (48 rows of 48 plants). Each plant has a different set of "RescueMu" mutations.

"We planned on starting the prototype field this summer at Stanford, but the El Niņo weather made it too iffy," Walbot says. "So we are going to do the first grid this winter in Molokai."

The researchers will record the development of each plant at the kernel, seedling, adult and flowering stages. Self-pollinating seeds will be saved from all the plants and placed in a government-funded seed bank, the Maize Genetics Cooperation Stock Center at the University of Illinois. Tissue samples will be taken from these plants. The 500,000-plus tagged genes that they contain will be removed and put into bacterial cultures, called libraries. There will be one library for every row and column in the corn field. Records will be kept associating each set of genes with the mutations expressed in the original plant.

By recovering individual "RescueMu" sequences from the libraries, the researchers plan to sequence and annotate more than 150,000 of these tagged genes using standard sequencing techniques. This should give them a 95 percent probability of having mapped all 50,000 genes that the scientists estimate make up the maize genome. The full collection will be distributed to other researchers to search for more genes.

"In the past, we have studied one gene at a time," says Walbot. "With our project, and the other plant genomic projects, we can begin looking at the effects of groups of genes. This should give us much deeper insight into the way that plants work as entire systems."

By identifying the function of natural corn genes, the maize geneticist also argues that the project should allow continued improvement in productivity without having to resort to inserting genes from other organisms to make controversial transgenic plant varieties.

An important aspect of the project is dissemination of the information that it produces. The researchers will develop web-based records and databases to allow other scientists to put their research to use. Bacterial culture grids, seeds and other materials will be maintained permanently by the University of Illinois center, which will distribute them to anyone interested at low cost.

"We wanted to make certain that there is no possible conflict of interest," says Walbot, who is a member of the board of directors of Pioneer Hi-Bred, the world's largest corn seed company. The other scientists involved also have close links with various agricultural companies.

Walbot's co-investigators are Sarah Hake and Michael Freeling from the University of California-Berkeley; Robert Schmidt and Laurie Smith from the University of California-San Diego; Vicki Chandler, Brian Larkins and David Galbriath from the University of Arizona; Marty Sachs from the University of Illinois; and Volker Brendel from Iowa State University.

The potential commercial value of the tools and information the project will generate is suggested by the fact that the National Corn Growers Associated lobbied hard for passage of the legislation that provided the money, and organized a national telephone teleconference on Monday to announce its establishment. Stanford's Office of Technology Licensing is organizing an industrial affiliates program for the project, similar to the technology transfer programs that have been established for a number of engineering centers and departments. The companies who sign up will have equal access to all the materials that the project produces. The cost of joining: $500,000 for the first year and $250,000 per year thereafter.

By David F. Salisbury

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