Analysis Software Images the Forgotten Crop.

Daniel S. Burgess

Alfalfa is the forgotten great crop in the US. As the fourth-largest in the country, behind corn, soybeans and wheat, the bulk of the estimated $6.4 billion annual alfalfa harvest is destined for animal feed as hay. Because little alfalfa makes its way directly to the dinner table, commercial researchers have overlooked it.

Unlike corn and soybeans, alfalfa is autotetraploid: The genes that determine the legume's traits are on four chromosomes instead of two. These chromosomes are particularly small -- on the order of 2 to 4 µm. The lack of gastronomic interest and the difficulty in mapping alfalfa's genes have meant little improvement of the plant; the average alfalfa yield has increased by only about 1 percent in the last 30 years.

Researchers at the US Department of Agriculture's Agricultural Research Service have found a solution to the mapping problem in an imaging system that they developed with Loats Associates Inc. The company's Inquiry software allows the group to magnify the chromosomes by up to 10,000 times and to color gray-scale images for banding studies, all on-screen.

Inquiry's smoothing algorithm and background correction enhance the image prior to quantitation. The combination of image processing and analysis enables the researchers to determine the shape and size of the plant's chromosomes and to locate the genes on them. An understanding of the gene sequence will follow, with the ability to selectively improve the crop.

"The image analysis system allowed us to increase the speed with which we could identify the individual chromosomes and thus create karyotypes," said Gary R. Bauchan, research geneticist at the service's Soybean and Alfalfa Research Laboratory. Before Inquiry, he said, "it was necessary to take pictures of the cells we were studying, develop the negative, print the pictures and take measurements off of the photomicrographs. This was a very laborious system."

An image analysis system from Loats Associates Inc. helps plant scientists at the Agricultural Research Service to investigate the genetics of alfalfa. The Inquiry software assigns false colors to the alfalfa chromosomes for banding studies and enables magnification of a single chromosome to 10,000×.

The technique has allowed the researchers to distinguish the chromosomes of the yellow-flowered Falcata subspecies of alfalfa from its purple-flowered, cultivated cousin. Falcata displays favorable traits, such as winter hardiness, persistence and resistance to some diseases, but its yellow blossoms are inefficient at attracting UV-sensitive bees for pollination.

The researchers have increased the resolution of the system's digital video camera to 2056 × 1920. "We have only begun to utilize this technology to identify chromosomes for the improvement of alfalfa," Bauchan said.