Update on SNP
The August/September 2010 issue of The Sunflower introduced readers to a topic that’s murky and mysterious to most of us — but one that is directly influencing the development of future sunflower hybrids. That topic is “SNP,” which stands for single-nucleotide polymorphism.
Before your eyes glaze over and your tongue fumbles while trying to digest that term, consider the implications of SNPs for sunflower producers and the industry in general.
SNPs are markers, tiny pieces of DNA located in or near a gene. Markers are indicators of an organism’s genetic makeup. In the crop world, as molecular biologists and plant breeders increase their ability to locate and identify markers for specific traits, they correspondingly become more effective at plugging the desired traits into new breeding populations and, ultimately, the commercial varieties planted by farmers.
“SNPs allow for detailed analysis of breeding populations using methods we’ve never used before,” noted Brent Hulke, USDA-ARS sunflower research geneticist, in that August/September article. Hulke used an analogy to further illustrate the importance of SNP technology:
“Up until now, markers have been pieces of DNA that, in most cases, are not associated directly with specific genes. They’re just ‘hooks.’ So it’s kind of like putting a stake next to a rock in a field. You can’t put that stake directly on the rock; but you can place it by the rock. . . . With SNPs, we’re going to a technology that can actually ‘put the stake right through the middle of the rock.’ We’re to the point now where we can easily get SNPs that are actually a part of the gene of interest.”
Why is it so important for the sunflower sector to pursue and develop SNP technology? In a word, competition. Other crops — corn prominent among them — have been employing SNP and doubled-haploid technology quite successfully. One glaring example is the imminent introduction of drought-tolerant corn varieties. Their availability in rainfall-short areas where sunflower has done well through the years could well translate into less sunflower acreage if sunflower is unable to compete agronomically and economically.
So while today’s sunflower hybrids certainly reflect significant breeding progress, there’s a need for even-faster progress if the industry is to remain competitive. The sunflower grower needs strong yields, good quality and an attractive price. SNP and doubled-haploid technology can help provide those first two components — which, in turn, obviously impact that all-important third one.
Recognizing the importance of this technology, last March the National Sunflower Association Board of Directors allocated $100,000 to help propel research aimed at facilitating marker-assisted breeding in sunflower. The first phase consisted of “sequencing,” i.e., measuring absolutely the DNA base composition of an organism. Phase One identified six sunflower breeding lines that together contained a lot of genetic diversity. Sequencing of those six lines, spearheaded by BioDiagnostics, Inc., a River Falls,Wis.-based independent DNA testing laboratory, was achieved in late August 2010.
The second phase is more involved and expensive. “Phase Two is the point at which we take the SNPs we’ve already discovered from our simple panel of six genotypes (inbreds) and try to see how common they are across a wide range of lines,” Hulke explains. Along with numerous lines from Hulke’s research program, Phase Two includes lines from the USDA-ARS Plant Introduction Station in Ames, Iowa, as well as lines submitted by commercial sunflower breeders. As of early December, about 1,300 lines were being analyzed.
“The goal is to find out which are the most informative markers,” Hulke relates. Informative markers are ones that show clear genetic differences among the 1,300 lines. The informative markers can then be focused on projects like finding resistance to Sclerotinia stalk rot — which, because it is a multi-gene trait, has always been a very vexing and difficult disease for which to find and incorporate resistance via traditional phenotypic selection. “But we have enough stalk rot resistance data on important lines to associate with our newly discovered SNP loci, once those become available,” Hulke says. The ARS geneticist believes “we will find many QTL (quantitative trait loci — stretches of DNA closely linked to the gene of interest) responsible for resistance, based on the diversity of the panel and the polygenic nature of Sclerotinia resistance.”
Phases Three and Four, for which Hulke is preparing a grant proposal to the National Sclerotinia Initiative, take on a fascinating new dimension. As soon as marker trait associations are finished for stalk rot, he proposes to work with a proprietary technology developed by California-based Illumina, Inc., to use those markers in a breeding program for their stalk rot resistance value. Illumina’s BeadXpress Reader System is essentially a form of nanotechnology, using bar-coded microbeads on a glass plate. Each bead is incredibly small — just 240 microns (one-fourth of one millimeter) in length. The plant DNA is placed on the beads, and the system then provides rapid and precise computer-based evaluation. That information allows breeders to know immediately whether a given line possesses the gene(s) of interest.
The information gleaned from this SNP program — which may be available to the sunflower breeding community within a year — will not be a panacea resulting in immediate and/or complete resistance to Sclerotinia stalk rot, Hulke emphasizes. But it will be an extremely valuable tool that he and other breeders can use in concert with the already-existing body of phenotypic data and ratings for Sclerotinia resistance.
What this will give sunflower breeders, the ARS geneticist says, is the ability to test for stalk rot resistance value right after they make the parental crosses. They will not have to wait until the hybrid development stage. “If your lines were genotyped with the 10,000 markers in Phase Two, you know which of those lines have the QTL for stalk rot,” Hulke illustrates. “If I say ‘SNP number 317 is highly significant for stalk rot resistance,’ the breeder can just review pedigree records and trace it back.” Bottom line: the breeder will know — up front, prior to any greenhouse or field testing — which parents would be suitable for making crosses to include Sclerotinia resistance.
Though Sclerotinia resistance likely will be the first sunflower horse out of the SNP gate, Hulke envisions eventually working on additional diseases, as well as other yield- and quality-influencing traits. “Anything we can measure in the field with repeatability is fair game,” he says.
The use of genotypic tools such as SNP is not going to replace phenotypic evaluation, Hulke emphasizes. “People tend to think that once we have the markers, that’s instantly superior to whatever data we can take from the field. That’s categorically false,” he states. “It’s an inferior replacement to field-derived data. But, it’s replacing during a phase of the breeding cycle where we had no information before. It’s a source of information that before was invisible.
“Nothing has changed genetically. We just have tools that help us understand the information better and to utilize it much quicker. Breeders will find they can be more efficient. They’re going to get more advancement within the same amount of time.”
Still, each seed company must do its own cost:benefit analysis, Hulke realizes. For instance, a company that relies almost exclusively on field testing and whose research budget is heavily weighted in that direction will need to decide whether diverting a portion of that budget toward SNP and doubled-haploid technologies is worthwhile.
For the sunflower producer, of course, it all comes down to being able to grow hybrids that are increasingly productive and profitable. If the sunflower seed company’s use of SNPs and doubled haploids helps achieve that — and does so in a significantly shorter time frame — it’s all good. — Don Lilleboe
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